KR20100095564A - Methods and compositions for assessing responsiveness of b-cell lymphoma to treatment with anti-cd40 antibodies - Google Patents

Abstract

The present invention provides methods and kits useful for predicting or evaluating the responsiveness of B-cell lymphoma to treatment with anti-CD40 antibodies.

Description

METHODS AND COMPOSITIONS FOR ASSESSING RESPONSIVENESS OF B-CELL LYMPHOMA TO TREATMENT WITH ANTI-CD40 ANTIBODIES}

Related application

This application claims priority based on US patent provisional application 60 / 986,277, filed November 7, 2007, which is incorporated by reference in its entirety.

FIELD OF THE INVENTION The present invention generally relates to the field of predicting, evaluating, or helping to assess the responsiveness of B-cell lymphoma to treatment with anti-CD40 antibodies.

CD40 is a type I transmembrane protein of the tumor necrosis receptor superfamily. CD40 is an important molecule involved in B-cell proliferation and differentiation, immunoglobulin isotype switching, and cell viability. Receptor signal transduction is initiated primarily by binding of CD40 to CD40 ligand (CD40L or CD154) expressed on activated CD4 + T cells.

On normal cells, CD40 is expressed on proliferative cells such as hematopoietic progenitor cells, epithelial and endothelial cells, and all antigen-presenting cells (dendritic cells, activated B lymphocytes, and activated monocytes). CD40 is highly expressed on several species of B-cell hematologic malignancies such as multiple myeloma, non-Hodgkin lymphoma (NHL), and chronic lymphocytic leukemia (CLL). The large preponderance of CD40 expression on B-cell malignancies is an attractive potential tumor target for antibody-based cancer therapy. CD40 is also expressed in most bladder cancers and in significant proportions of other solid tumors such as head and neck cancer, renal cell carcinoma, ovarian cancer and lung cancer.

Anti-CD40 antibodies, and their use for treating B cell hematologic malignancies, have been described (eg, US Pat. No. 6,946,129; 6,843,989; 6,838,261; WO 2000/075348; US-2002-0197256; US Pat. No. 2006/128103; and See WO 2007/075326). Humanized anti-CD40 antibodies have been shown to induce growth inhibition and apoptosis of CD40-positive cells in a subset of blood tumor cell lines via direct signal transduction (WO 2006/128103; WO 2007/075326). Humanized anti-CD40 antibodies also kill tumor cells through immune effector functions such as antibody dependent cellular cytotoxicity (ADCC) and antibody dependent cellular phagocytosis (ADCP). In vivo, when using xenograft models of multiple myeloma (MM) and non-Hodgkin's lymphoma (NHL), anti-CD40 antibodies inhibit tumor growth and improve survival in severe combined immunodeficiency (SCID) mice. In some models, the anti-tumor activity of anti-CD40 antibodies has been found to be at least as effective as Rituximab compared to Rituximab (Genentech, Inc.).

Phase I clinical trials with humanized anti-CD40 antibodies in 2004 at Seattle Genetics were initiated as a single agent multi-dose trial in patients with recurrent and refractory multiple myeloma (MM). Subsequently, phase I trials were initiated in patients with recurrent non-Hodgkin's lymphoma (NHL) and chronic lymphocytic lymphoma (CLL). Results from these phase I trials showed evidence of anti-tumor activity in patients with myeloma with stable disease and reduced M-protein, NHL patients with partial and complete response, and CLL patients with stable disease. Phase II testing of anti-CD40 antibodies in recurrent diffuse large cell B cell lymphoma (DLBCL) was started in December 2006.

Although anti-CD40 antibodies have been shown to induce growth inhibition and apoptosis of CD40-positive cells and have anti-tumor activity in patients with various B cell lymphomas, all B lymphoma cells have anti-CD40 antibodies. It is not susceptible to mediated cell death. It is necessary to identify one or more predictive markers for the responsiveness of B-cell lymphoma patients to anti-CD40 antibody therapy.

All references cited herein, including patent applications and publications, are incorporated herein by reference in their entirety.

Summary of the Invention

The present invention provides methods and compositions for predicting, evaluating, or assisting in the assessment of a subject with a kind of B-cell lymphoma to treatment with an anti-CD40 antibody.

In one aspect, the invention includes comparing the measured expression level of at least one marker gene in any of Tables 2-4, 6, 7, and 13 to a reference level in a B-cell lymphoma sample from a subject Methods are provided to assess or assist in the responsiveness of a subject with B-cell lymphoma to treatment with a CD40 antibody.

In another aspect, the invention includes comparing the measured expression level of at least one marker gene in any of Tables 2-4, 6, 7, and 13 to a reference level in a B-cell lymphoma sample from a subject Provides methods for predicting responsiveness to anti-CD40 antibody treatment or monitoring treatment / responsiveness in subjects with cell lymphoma.

In another aspect, the invention measures (a) the expression level of one or more marker genes in a sample comprising B lymphoma cells obtained from a subject with B-cell lymphoma, wherein the one or more marker genes are selected from IFITM1, CD40, RGS13. , VNN2, LMO2, CD79B, CD22, BTG2, IGF1R, CD44, CTSC, EPDR1, UAP1, and PUS7; (b) predicting whether the subject will respond to anti-CD40 antibody treatment based on the measured expression level of the one or more marker genes from step (a). Methods are provided for predicting, evaluating, or assisting in the assessment of a subject with cellular lymphoma. In some embodiments, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, or 14 marker genes from said group. Expression levels are measured and used for prediction or evaluation or to aid evaluation. In some embodiments, helping to predict or assess or assess is determined by comparing the measured expression levels of one or more marker genes to a reference level. In some embodiments, the reference level is a value or range determined based on the measured expression level of the corresponding marker gene in a sample comprising B lymphoma cells from a subject having increased or decreased tumor volume after anti-CD40 antibody treatment. .

In another aspect, the invention provides an antibody comprising (a) IFITM1, CD40, RGS13, VNN2, LMO2, CD79B, CD22, BTG2, IGF1R, CD44, CTSC, EPDR1 in a sample comprising B lymphoma cells from a subject having B-cell lymphoma. Determining the expression level of one or more marker genes selected from the group consisting of UAP1, PUS7, and BCL6; (b) providing a method of preparing a personalized genomics profile for a subject with B-cell lymphoma, comprising preparing a report summarizing the expression levels of one or more marker genes obtained in step (a) do. In some embodiments, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, or 15 from said group. The expression level of the marker gene is measured and used to report on a personalized genomics profile. In some embodiments, the report includes recommendations for anti-CD40 antibody treatment for the subject. In some embodiments, the recommendation is determined by comparing the measured expression level of the marker gene to the reference level. In some embodiments, the reference level is a value or range determined based on the measured expression level of the corresponding marker gene in a sample comprising B lymphoma cells from a subject having increased or decreased tumor volume after anti-CD40 antibody treatment. .

In another aspect, the invention provides an antibody comprising (a) IFITM1, CD40, RGS13, VNN2, LMO2, CD79B, CD22, BTG2, IGF1R, CD44, CTSC, EPDR1 in a sample comprising B lymphoma cells from a subject with B-cell lymphoma. Measuring expression levels of at least two marker genes selected from the group consisting of UAP1, and PUS7; (b) calculating the sensitivity index (SI) value based on the measured expression level of the marker gene in step (a) by the following formula: B-cells for anti-CD40 antibody treatment Provides methods for predicting, evaluating, or assisting in evaluating the responsiveness of a subject with lymphoma:

Wherein the expression levels of at least one marker gene having a positive correlation value and at least one marker gene having a negative correlation value are determined;

(i) β _j is the coefficient value for each marker gene measured; (ii) p is the number of marker genes measured; (iii) χ _i is a normalized expression level, transformed for a sample from the subject for the expression level of each marker measured; (iv) μ _j and σ _j are the mean and standard deviation for each marker gene measured; Wherein β _j , μ _j and σ _j are determined from a patient sample comprising B lymphoma cells. In some embodiments, a value above zero for the sensitivity index indicates whether the subject will respond to anti-CD40 antibody treatment, or a value below zero for the sensitivity index indicates whether the subject will respond less to anti-CD40 antibody treatment. Indicates. In some embodiments, the expression level of at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, or 14 marker genes is measured, Used to calculate sensitivity index. In some embodiments, the expression levels of IFITM1, RGS13, CD79B, CD22, BTG2, CD44, EPDR1, and UAP1 are measured and used for calculating the sensitivity index.

In another aspect, the present invention provides a method of treating a subject with B-cell lymphoma, comprising administering to the subject an effective amount of an anti-CD40 antibody, wherein responsiveness of the B-cell lymphoma in the subject is described herein. It was evaluated by the method. In another aspect, the invention provides a) at least of any of Tables 2-4, 6, 7 and 13 in a B-cell lymphoma sample from a subject to assess whether the subject's B-cell lymphoma is suitable for anti-CD40 antibody treatment. Selecting a subject for anti-CD40 antibody treatment by comparing the measured expression level of one marker gene to a reference level; A method of treating a subject with B-cell lymphoma, comprising administering to the subject an effective amount of an anti-CD40 antibody.

In some embodiments, the reference level is the measured expression level of one or more reference genes of Table 8 or Table 9 in a B-cell lymphoma sample from the subject.

In some embodiments, the reference level is a measured expression level of a marker gene in different B-cell lymphoma samples. In some embodiments, different B cell lymphoma samples comprise B lymphoma cells that are resistant to anti-CD40 antibody induced cell death.

In some embodiments, the measured expression level and / or reference level of the marker gene is normalized.

In some embodiments, measured expression levels of at least 2, at least 5, at least 10, at least 15, or at least 20 genes in any of Tables 2-4, 6, 7, and 13 in a B-cell lymphoma sample from a subject. Compare to one or more reference levels.

In some embodiments, expression levels are measured by detecting mRNA expression (eg, real time quantitative reverse transcription PCR (qRT-PCR)) and / or by detecting protein expression (eg, immunohistochemistry (IHC)). .

In some embodiments, the marker genes measured include one or more CD40 ligand downregulated genes (eg, VNN2, MEF2C, LTB, KCNN3, NCF1, BCL6, IGJ, ELTI1902, PNOC, CSF2RB, and POU2AF1). In some embodiments, the marker genes measured comprise one or more genes (eg, CD22, RGS13, and MEF2B) in the B-cell receptor signal transduction pathway.

In some embodiments, at least one selected from the group consisting of VNN2, MEF2C, LTB, KCNN3, NCF1, BCL6, IGJ, ELTI1902, PNOC, CSF2RB, POU2AF1, CD22, RGS13, and MEF2B in a B-cell lymphoma sample from a subject At least one, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, or 14 genes. Compare.

In some embodiments, at least one selected from the group consisting of VNN2 and EPDR1, RGS13 and EPDR1, CD22 and EPDR1, LRRC8A and PRPSAP2, CD40 and IGF1R, IFITM1 and BTG2, SMN1 and LMO2, PRKCA and YIPF3 in B-cell lymphoma samples Compare the expression level of the gene pairs. In some embodiments, the expression level between one or more gene pairs VNN2 and EPDR1, RGS13 and EPDR1, CD22 and EPDR1, LRRC8A and PRPSAP2, CD40 and IGF1R, IFITM1 and BTG2, SMN1 and LMO2, PRKCA and YIPF3 in a B-cell lymphoma sample To evaluate the responsiveness of B-cell lymphoma to anti-CD40 antibody treatment by calculating the sensitivity index calculated as the sum of the signed t-scores for log2-scale expression of the gene pair. Used for.

In some embodiments, the B-cell lymphoma is non-Hodgkin's lymphoma (NHL), including but not limited to follicular lymphoma, recurrent follicular lymphoma, small lymphocytic lymphoma, mantle cell lymphoma, marginal lymphoma, lymphoid cell Lymphoma, mycelial sarcoma / Sezary syndrome, splenic marginal lymphoma, and diffuse large cell B-cell lymphoma. In some embodiments, the B-cell lymphoma is selected from the group consisting of indolent lymphomas, aggressive lymphomas, and highly aggressive lymphomas.

In a further aspect, the present invention provides a kit comprising a reagent for measuring the expression level of at least one marker gene of any of Tables 2-4, 6, 7, and 13. In some embodiments, the kit comprises at least one pair of primers for amplifying at least one marker gene of any of Tables 2-4, 6, 7, and 13 by PCR. For example, the forward and reverse primers shown in Table 10 can be used. The kit may further comprise a pair of primers for amplifying the reference genes of Table 8. The kit may further comprise a surface to which a probe for detecting the amplified gene product is attached, for example a microarray, in which the present invention contemplates and includes such a surface. In some embodiments, the kit comprises at least one pair of primers and probes for detecting the expression level of one marker gene in any of Tables 2-4, 6, 7, and 13 by qRT-PCR. The kit may further comprise a pair of primers and probes for detecting the expression level of the reference genes of Table 8 by qRT-PCR. For example, the primer and probe sets shown in Table 10 can be used for the detection of expression levels of genes by qRT-PCR. In some embodiments, the kit comprises one or more antibodies that specifically recognize one or more proteins encoded by the marker gene. The kit may further comprise instructions for carrying out other reagents and / or any of the methods described herein.

It should be understood that one, some or all of the features of the various embodiments described herein may be combined to form other embodiments of the invention. These and other aspects of the invention will be apparent to those skilled in the art.

Enrichment plots of genes in the BASSO_GERMINAL_CENTER_CD40_DN gene set. The top plot shows enrichment score distribution across genes ranked from the modulated t-test (Table 2). The lower plot shows the distribution of enrichment on a ranked list metric known as signal2noice. Taken together, these plots clearly show that the gene set is strongly enriched in anti-CD40 Ab.1 sensitive cells.
Figure 2. VNN2 (CD40L-downregulated gene) is overexpressed in sensitive NHL cells for anti-CD40 Ab.1 and distinguishes between two classes of sensitivity and resistance. Bar graphs show mRNA expression levels and line graphs show IC25 values.
Figures 3A-3C. RGS13, CD22, and MEF2B germinal center B markers are overexpressed in sensitive and intermediate NHL cells for anti-CD40 Ab.1 and distinguish with reasonable accuracy between the two classes of sensitivity and resistance can do. Bar graphs show mRNA expression levels and line graphs show IC25 values.
4. Anti-CD40Ab.1 sensitivity index scoring across NHL cell lines. Stepwise Linear Modeling and gene-pair scoring were applied to each cell line based on mRNA expression data. The primary y-axis shows anti-CD40 Ab.1 sensitivity index and the secondary y-axis shows anti-CD40 Ab.1 IC25 values plotted against NHL cell lines on the x-axis. High anti-CD40 Ab.1 sensitivity index (> -4) indicates an increased likelihood that the cell line is sensitive.
5. Correlation of anti-CD40.Ab.1 susceptibility and CD40 signature genes.
Figures 6a-6ii. Gene Bank sequences for the genes listed in Tables 7 and 10. VNN2 (FIG. 6A: SEQ ID NO: 258), RGS13 (FIG. 6B: SEQ ID NO: 259), CD22 (FIG. 6C and 6D: SEQ ID NO: 260), LRRC8A (FIG. 6E: SEQ ID NO: 261), CD40 (FIG. 6F: SEQ ID NO: 262), IFITM1 (FIG. 6g: SEQ ID NO: 263), PRKCA (FIGS. 6H-6J: SEQ ID NO: 264), BCL6 (FIGS. 6K and 6L: SEQ ID NO: 265), EPDR1 (FIG. 6M: SEQ ID NO: 266), PRPSAP2 (FIG. 6N: SEQ ID NO: 267), IGF1R (FIG. 6O To 6r: SEQ ID NO: 268), BTG2 (FIG. 6s and 6t: SEQ ID NO: 269), LMO2 (FIG. 6u: SEQ ID NO: 270), YIPF3 (FIG. 6V: SEQ ID NO: 271), SMN1 (FIG. 6W: SEQ ID NO: 272), CD79B (FIG. 6x: SEQ ID NO: 273), CD44 (FIGS. 6Y and 6z: SEQ ID NO: 274), CTSC (FIG. 6aa: SEQ ID NO: 275), UAP1 (FIG. 6BB: SEQ ID NO: 276), PUS7 (FIG. 6CC and 6dd: SEQ ID NO: 277), RGS13 (FIG. 6EE: Sequence) 278), nucleic acid sequences encoding mRNAs of CD22 (FIGS. 6ff and 6gg: SEQ ID NO: 279), SMN1 (FIG. 6hh: SEQ ID NO: 280), and YIPF3 (FIG. 6ii: SEQ ID NO: 281).
7. Correlation of the percent change in multivariate susceptibility index and sum of product of diameter of tumor (SPD) measurements for 21 patients in clinical trial 001. The percent SPD change is determined by comparing the minimum post-baseline SPD to the baseline SPD. Positive changes indicate an increase in tumor volume and negative changes indicate a decrease in tumor volume. The weights (coefficients) used to calculate the sensitivity index are shown in Table 14. Larger multivariate sensitivity index values are associated with post-baseline SPD reduction (Spearman Rho = -0.58; P = 0.006).
8. Association of% change in BCL6 expression and SPD measurements for 26 DLBCL patients. The percent SPD change is determined by comparing the minimum post-baseline SPD to the baseline SPD. Positive changes indicate an increase in tumor volume and negative changes indicate a decrease in tumor volume.

The present invention is resistant to B lymphoma cells and anti-CD40 antibody induced cell death in which certain genes (eg, those shown in Tables 2-4, 6, 7, and 13) are susceptible to anti-CD40 antibody induced cell death. It is based on the discovery that it is differentially expressed between phosphorus B lymphoma cells. The clinical trial data described in Example 2 indicate that the expression levels of the 14 genes shown in Table 13 are highly associated with responsiveness to anti-CD40 Ab.1 treatment. Some of the genes differentially expressed between sensitive B lymphoma cells and resistant B lymphoma cells are CD40 ligand downregulated pathway genes; Some are within the B-cell receptor signal transduction pathway. Thus, the expression level of one or more of these differentially expressed genes may be used to assess or assist in the responsiveness of a subject with B-cell lymphoma to treatment with an anti-CD40 antibody, and to treat with an anti-CD40 antibody. It can be used to predict the subject's responsiveness to the subject and to monitor treatment / responsiveness in the subject.

A. General Skills

The practice of the present invention will employ, unless otherwise indicated, conventional techniques of molecular biology (including recombination techniques), microbiology, cell biology, biochemistry and immunology, which are within the skill of the art. Such techniques are described in Molecular Cloning: A Laboratory Manual, second edition (Sambrook et al., 1989); Oligonucleotide Synthesis (M. J. Gait, ed., 1984); Animal Cell Culture (R.I. Freshney, ed., 1987); Methods in Enzymology (Academic Press, Inc.); Current Protocols in Molecular Biology (F.M. Ausubel et al., Eds 1987, and periodic updates); It is fully described in literature such as PCR: The Polymerase Chain Reaction (Mullis et al., Ed., 1994).

Primers, oligonucleotides and polynucleotides used in the present invention can be produced using standard techniques known in the art.

Unless defined otherwise, technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Singleton et al., Dictionary of Microbiology and Molecular Biology 2nd ed., J. Wiley & Sons (New York, NY 1994) and March, Advanced Organic Chemistry Reactions, Mechanisms and Structure 4th ed., John Wiley & Sons (New York, NY 1992) provides general guidance for many terms used herein to those skilled in the art.

B. Definition

As used herein, the terms "subject with B-cell lymphoma" and "patient with B-cell lymphoma" refer to a subject diagnosed with or capable of being diagnosed with a type of B-cell lymphoma.

The term "biomarker" or "marker", as used herein, can be detected by known methods (or methods disclosed herein) and its expression or secretion in or on mammalian tissues or cells and anti-CD40 antibodies To predict (or assist in) predicting the susceptibility of a mammalian cell or tissue to a therapeutic regimen based on, and in some embodiments, to predict an individual's responsiveness to a therapeutic regimen based on an anti-CD40 antibody ( Or molecules comprising genes, proteins, carbohydrate structures, or glycolipids, which can be used).

The term "sample", as used herein, contains cellular and / or other molecular entities that are characterized and / or identified, eg, based on physical, biochemical, chemical, and / or physiological characteristics, By a composition is obtained or derived from the desired subject. For example, the phrase “disease sample” and its modified expression means any sample obtained from a subject of interest that is expected to contain, or known to contain, the cellular and / or molecules being characterized.

"Tissue or cell sample" means a collection of similar cells obtained from a tissue of a subject or patient. The source of tissue or cell sample may be solid tissue, such as from fresh, frozen and / or preserved organ or tissue sample or biopsy or aspirate; Blood or any blood construct; Body fluids such as cerebrospinal fluid, amniotic fluid, peritoneal fluid, or interstitial fluid; Cells from any time during pregnancy or development of the subject. In addition, the tissue sample may be a primary or cultured cell or cell line. Optionally, tissue or cell samples are obtained from diseased tissue / organs. Tissue samples may contain compounds that do not naturally mix with the tissue in nature, such as preservatives, anticoagulants, buffers, fixatives, nutrients, antibiotics, and the like.

For the purposes herein, "fragment" of a tissue sample means a single portion or piece of tissue sample, eg, a thin piece of tissue or cells cut from a tissue sample. If it is understood that the present invention includes a method in which the same fragment of a tissue sample is analyzed at both morphological and molecular levels, or for both protein and nucleic acid, several sections of the tissue sample may be taken and analyzed for analysis according to the present invention. It will be appreciated that it can be applied.

As used herein, a "B-cell lymphoma sample" or "sample comprising B lymphoma cells" is a tissue or cell sample containing B lymphoma cells obtained from a subject or patient diagnosed with a kind of B-cell lymphoma.

As used herein, a method for "helping with evaluation" means a method that helps clinical decisions (eg, responsiveness of B-cell lymphoma to treatment with an anti-CD40 antibody) and may be critical for final evaluation. It may or may not be.

"Subject" or "subject" is a mammal, more preferably a human. Mammals include, but are not limited to, humans, primates, livestock, sport animals, rodents, and pets (eg, dogs and cats).

As used herein, a "reference value" means an absolute value; Relative value; Values having an upper limit and / or a lower limit; Range of values; medium; Median value; Median value; Or a value compared to a particular control or baseline value.

The term "array" or "microarray" as used herein refers to a regular alignment on a substrate of hybridizable array elements, eg, polynucleotide probes (eg oligonucleotides) and antibodies. The substrate may be a solid substrate, for example a glass slide, or a semisolid substrate, for example a nitrocellulose membrane. The nucleotide sequence can be DNA, RNA, or any variant thereof.

"Amplification", as used herein, generally refers to the process of producing multiple copies of a sequence of interest. "Multiple copies" means at least two copies. "Copy" does not necessarily mean complete sequence complementarity or identity to the template sequence. For example, the copy may be a nucleotide analogue such as deoxyinosine, intentional sequence alteration (eg, a sequence alteration introduced through a primer comprising a sequence that is hybridizable to a template but is not complementary), and / or Sequence errors that occur during amplification.

The expression / amount of the gene or biomarker in the first sample is at least about 1.5X, 1.75X, 2X, the expression level / amount of the gene or biomarker in the first sample is at least about 1.5X, 1.75X, 2X, 3X, 4X, 5X, 6X, 7X, 8X, 9X or 10X is a level "greater than" in the second sample. Expression levels / amounts can be determined based on any suitable criteria known in the art, including but not limited to mRNA, cDNA, proteins, protein fragments and / or gene copies. Expression levels / amounts can be determined qualitatively and / or quantitatively.

"Polynucleotide" or "nucleic acid" as used herein interchangeably means a polymer of nucleotides of any length and includes DNA and RNA. Nucleotides can be deoxyribonucleotides, ribonucleotides, modified nucleotides or bases, and / or their analogs, or any substrate that can be incorporated into a polymer by DNA or RNA polymerase. Polynucleotides may include modified nucleotides such as methylated nucleotides and their analogs. If present, modifications to the nucleotide structure can be imparted before or after assembly of the polymer. The sequence of nucleotides may be interrupted by non-nucleotide components. The polynucleotides may be further modified after polymerization, for example by conjugation of the labeling component. Other kinds of modifications include, for example, "cap" substitutions of one or more naturally occurring nucleotides with analogues, internucleotide modifications, eg, non-charged linkages (eg, methyl phosphonates, phosphorides). Modifications with esters, phosphoramidates, carbamates, and the like, and with charged groups (eg, phosphorothioates, phosphorodithioates, etc.), suspended moieties such as proteins (eg, For example, those containing nucleases, toxins, antibodies, signal peptides, poly-L-lysine, etc., those having an intercalator (eg, acridine, solarene, etc.), chelators containing chelators (e.g., metals, radioactive metals, boron, metal oxides, etc.), containing alkylators, modified linkages (e.g., alpha anomeric nucleic acids, etc.) Of the polynucleotide (s) And a modified form. In addition, any hydroxyl group normally present in the sugar may be replaced by, for example, a phosphonate group, a phosphate group, protected with a standard protecting group, or activated to prepare additional linking groups for additional nucleotides, It may be conjugated to a solid or semi-solid support. 5 'and 3' terminal OH can be phosphorylated or substituted with amines or organic capping group moieties of 1 to 20 carbon atoms. Other hydroxyls may also be derivatized to standard protecting groups. Polynucleotides are also generally analogous to ribose or deoxyribose sugars known in the art, for example 2'-0-methyl-2'-0-allyl, 2'-fluoro- or 2'-azido -Ribose, carbocyclic sugar analogs, α-anomeric sugars, epimer sugars such as arabinose, xylose or lysose, pyranose sugar, furanos sugar, sedoheptulose, acyclic analogues and nonbasic nucleoles Seed analogs such as methyl riboside. One or more phosphodiester linkers may be replaced with replacement linkers. These alternative linkers include phosphates having P (O) S ("thioate"), P (S) S ("dithioate"), (O) NR ₂ ("amidate"), P (O) R, P (O) OR ', CO or CH ₂ ("formacetal"), wherein each R or R' is independently H or optionally substituted or unsubstituted alkyl (1-20) containing an ether (-O-) linking group C), aryl, alkenyl, cycloalkyl, cycloalkenyl, or araldyl). Not all linking groups in the polynucleotide need to be identical. The above description applies to all polynucleotides referred to herein, including RNA and DNA.

As used herein, “oligonucleotide” refers to a short, generally single-stranded, generally synthetic polynucleotide that is not necessarily, but generally less than about 200 nucleotides in length. The terms "oligonucleotide" and "polynucleotide" are not intended to exclude each other. The above description of polynucleotides is equally applicable to oligonucleotides.

A “primer” is a short single-stranded poly generally having free 3′-OH groups that binds to a target potentially present in the desired sample by hybridizing to the target sequence and then promotes polymerization of polynucleotides complementary to the target. Nucleotides. "Pair of primers" means 5 'primers and 3' primers that can be used to amplify a portion of a particular target gene.

The term “3 ′” generally refers to a region or location within a polynucleotide or oligonucleotide located 3 ′ (downstream) from another region or location within the same polynucleotide or oligonucleotide. The term "5 '" generally means a region or location within a polynucleotide or oligonucleotide located 5' (upstream) from another region or location within the same polynucleotide or oligonucleotide.

The phrase “gene amplification” refers to the process by which multiple copies of a gene or gene fragment are formed in a particular cell or cell line. Replicated regions (stretch of amplified DNA) are often referred to as "amplicons". In general, the amount of messenger RNA (mRNA) produced, ie the level of gene expression, also increases in proportion to the number of copies of the particular gene expressed.

“Detect” includes any detection means, including direct and indirect detection.

The term "prediction" refers herein to the likelihood that a patient will respond favorably or adversely to a drug or set of drugs. In one embodiment, the prediction is related to the degree of the response. In one embodiment, the prediction relates to whether and / or the likelihood that the patient will survive or ameliorate for a certain period of time without recurrence of the disease, for example after treatment with a particular therapeutic agent. The prediction method of the present invention can be used clinically to determine treatment by selecting the most appropriate treatment regimen for any particular patient. The predictive method of the present invention is intended to provide a patient with a favorable response to a treatment regimen, eg, a given treatment regimen, eg, administration of a given therapeutic agent or combination, surgical intervention, steroid treatment, or the like, or after a treatment regimen. Is a valuable tool for predicting the possibility of long-term survival.

The term “long term” survival is used herein to refer to survival for at least 1 year, 5 years, 8 years, or 10 years after treatment.

"Patient response" includes (1) inhibition of some degree of disease progression, including delayed and complete arrest; (2) reduction in the number of disease episodes and / or symptoms; (3) reduction of lesion size; (4) inhibition (ie, reduction, delay or complete stop) of disease cell infiltration into adjacent peripheral organs and / or tissues; (5) inhibition (ie, reduction, delay or complete stopping) of disease spread; (6) the reduction of some degree of one or more symptoms associated with the disease; (7) an increase in the period of time without disease after treatment; And / or (8) any endpoint indicating benefit for the patient, including but not limited to a reduction in mortality at any point after treatment.

The term “antibody” is used in its broadest sense and specifically refers to monoclonal antibodies (including full length monoclonal antibodies), multispecific antibodies (eg bispecific antibodies) and antibodies that exhibit the desired biological activity or function. Contains fragments.

An "antibody fragment" comprises a portion of a full length antibody, generally an antigen binding or variable region. Examples of antibody fragments include Fab, Fab ', F (ab') ₂ , and Fv fragments, diabodies; Linear antibodies; Single chain antibody molecules; And multispecific antibodies formed from antibody fragments.

"Fv" is the minimum antibody fragment that contains a complete antigen recognition and antigen binding site. This fragment consists of a dimer of one heavy chain and one light chain variable domain that are tightly noncovalently associated. Folding of the two domains results in six hypervariable loops (three loops each derived from H and L chains) that present amino acid residues for antigen binding and confer antigen binding specificity for the antibody. However, even a single variable domain (or half of the Fv containing only three CDRs specific for the antigen) has a lower affinity than the entire binding site but the ability to recognize and bind the antigen.

As used herein, the term “monoclonal antibody” refers to an antibody obtained from a substantially homogeneous population of antibodies, ie the production of monoclonal antibodies in which the individual antibodies that make up this population may generally be present in small amounts. Bind to the same and / or identical epitope (s) except for possible mutations that may occur during. The monoclonal antibody generally comprises an antibody comprising a polypeptide sequence that binds a target, and the target binding polypeptide sequence is obtained by a process comprising the selection of a single target binding polypeptide sequence from a plurality of polypeptide sequences. For example, the selection process may be the selection of unique clones from a number of clones, such as hybridoma clones, phage clones, or pools of recombinant DNA clones. Selected target binding sequences can, for example, improve affinity for a target, humanize target binding sequences, improve their production in cell culture, reduce their immunogenicity in vivo, and produce multispecific antibodies. It is to be understood that antibodies which may be further altered for purposes of this invention and which comprise the altered target binding sequence are also monoclonal antibodies of the invention. In contrast to polyclonal antibody preparations that generally include different antibodies directed against different determinants (epitopes), each monoclonal antibody of a monoclonal antibody preparation acts against a single determinant on an antigen. In addition to its specificity, monoclonal antibody preparations are generally advantageous in that they are not contaminated by other immunoglobulins. The alteration expression “monoclonal” refers to the properties of the antibody obtained from a substantially homogeneous population of antibodies and should not be considered as requiring antibody preparation by any particular method. For example, monoclonal antibodies used in accordance with the present invention may be used, for example, in hybridoma methods (eg, Kohler et al., Nature, 256: 495 (1975); Harlow et al., Antibodies). : A Laboratory Manual. (Cold Spring Harbor Laboratory Press, 2 ^nd ed. 1988); Hammerling et al., In: Monoclonal Antibodies and T-Cell Hybridomas 563-681 (Elsevier, NY, 1981)), recombinant DNA methods (See, eg, US Pat. No. 4,816,567), phage display technology (e.g., Clackson et al., Nature, 352: 624-628 (1991); Marks et al., J Mol. Biol. 222: 581-597 (1991); Sidhu et al., J Mol. Biol. 338 (2): 299-310 (2004); Lee et al., J Mol. Biol. 340 (5): 1073- 1093 (2004); Fellouse, Proc. Natl. Acad. Sci. USA 101 (34): 12467-12472 (2004); and Lee et al., J Immunol.Methods 284 (1-2): 119 -132 (2004)), and humans in animals having some or all of the genes encoding human immunoglobulin locus or human immunoglobulin sequence Techniques for producing human-like antibodies (eg, WO98 / 24893; WO96 / 34096; WO96 / 33735; WO91 / 10741; Jakobovits et al., Proc. Natl. Acad. Sci. USA 90: 2551 (1993) Jakobovits et al., Nature 362: 255-258 (1993); Brugemann et al., Year in Immunol. 7:33 (1993); U.S. Patent 5,545,806; 5,569,825; 5,591,669 (both GenPharm); 5,545,807; WO 1997/17852; U.S. Patent 5,545,807; 5,545,806; 5,569,825; 5,625,126; 5,633,425; And 5,661,016; Marks et al., Bio / Technology 10: 779-783 (1992); Lonberg et al., Nature 368: 856-859 (1994); Morrison, Nature 368: 812-813 (1994); Fishwild et al., Nature Biotechnology 14: 845-851 (1996); Neuberger, Nature Biotechnology 14: 826 (1996) and Lonberg and Huszar, Intern. Rev. Immunol. 13: 65-93 (1995)).

Monoclonal antibodies specifically include "chimera" antibodies herein. A “chimeric” antibody (immunoglobulin) is the same or homologous to the corresponding sequence of an antibody in which some of its heavy and / or light chains are from a particular species or belong to a particular antibody class or subclass, and the rest of the chain (s) Fragments of such antibodies that are identical to the corresponding sequence of an antibody from another species or belong to another antibody class or subclass and exhibit the desired biological activity (US Pat. No. 4,816,567; and Morrison et al. Proc. Natl. Acad. Sci. USA 81: 6851-6855 (1984)]. Humanized antibodies are a subset of chimeric antibodies as used herein.

A “humanized” form of a non-human (eg murine) antibody is a chimeric antibody comprising a minimal sequence derived from a non-human immunoglobulin. In most cases, a humanized antibody is a hypervariable region residue of a non-human species (donor antibody), for example mouse, rat, rabbit or non-human primate, in which the hypervariable region residues of the recipient have the desired specificity, affinity and ability. Substituted human immunoglobulins (donating or receiving antibodies). In some cases, framework region (FR) residues of human immunoglobulins are substituted with corresponding non-human residues. Humanized antibodies may also include residues that are not found in the donor antibody or donor antibody. Such modifications are intended to further improve antibody performance, eg binding affinity. In general, a humanized antibody will comprise substantially all of at least one, generally two variable domains, where all or substantially all hypervariable loops correspond to hypervariable loops of non-human immunoglobulins and all or substantially all FRs. The region is an FR region of the human immunoglobulin sequence, but the FR region may comprise one or more amino acid substitutions that improve binding affinity. The number of said amino acid substitutions in the FR is generally no more than 6 in the H chain and no more than 3 in the L chain. In addition, the humanized antibodies will optionally comprise an immunoglobulin constant region (Fc), generally at least a portion of the constant region of human immunoglobulin. For more details, see Jones et al., Nature 321: 522-525 (1986); Riechmann et al., Nature 332: 323-329 (1988); And Presta, Curr. Op. Struct. Biol. 2: 593-596 (1992).

A “human antibody” is an antibody comprising an amino acid sequence corresponding to an antibody produced by a human and / or prepared using any known human antibody preparation technique. The above definition of human antibody specifically excludes humanized antibodies comprising non-human antigen-binding residues.

An "affinity-promoting" antibody is an antibody that has one or more alterations in one or more CDR / HVRs thereof, which improves the affinity of the antibody for antigens relative to the parent antibody without alteration (s). Preferred affinity enhancing antibodies will have nanomolar or even picomolar affinities for the target antigen. Affinity-promoting antibodies are prepared by procedures known in the art. Marks et al., Bio / Technology 10: 779-783 (1992) describe affinity enhancement by VH and VL domain shuffling. Random mutagenesis of CDR / HVR and / or framework residues is described by: Barbas et al., Proc Nat. Acad. Sci. USA 91: 3809-3813 (1994); Chier et al., Gene 169: 147-155 (1995); Yelton et al., J. Immunol. 155: 1994-2004 (1995); Jackson et al., J. Immunol. 154 (7): 3310-9 (1995); And Hawkins et al., J. Mol. Biol., 226: 889-896 (1992).

The term “Fc region” is used to define the C-terminal region of an immunoglobulin heavy chain that can be produced by papain digestion of an intact antibody. The Fc region may be a native sequence Fc region or variant Fc region. While the boundaries of the Fc region of an immunoglobulin heavy chain can vary, the human IgG heavy chain Fc region is generally defined to extend from the amino acid residue at approximately position Cys226, or approximately position Pro230 to the carboxyl-terminus of the Fc region. The Fc region of an immunoglobulin generally comprises two constant domains, namely a CH2 domain and a CH3 domain, optionally comprising a CH4 domain. "Fc region chain" means herein one of the two polypeptide chains of an Fc region.

Antibody “effector function” means biological activity attributable to the Fc region (natural sequence Fc region or amino acid sequence variant Fc region) of an antibody, and differs depending on the antibody isotype. Examples of antibody effector functions include C1q binding and complement dependent cytotoxicity; Fc receptor binding; Antibody dependent cell mediated cytotoxicity (ADCC); Phagocytosis; Down regulation of cell surface receptors (eg B cell receptor); And B cell activation.

“Antibody dependent cell mediated cytotoxicity” or “ADCC” refers to the secreted Ig bound to the Fc receptor (FcR) present in certain cytotoxic cells (eg, natural killer (NK) cells, neutrophils and macrophages), By a cytotoxic effector cell specifically binds to an antigen bearing target cell, it refers to a form of cytotoxicity that causes the target cell to die with a cytotoxin. Antibodies "arm" cytotoxic cells and are absolutely necessary for such killing. NK cells, the primary cells that mediate ADCC, express only FcγRIII and monocytes express FcγRI, FcγRII, and FcγRIII. FcR expression on hematopoietic cells is described by Ravetch and Kinet, Annu. Rev. Immunol., 9: 457-92 (1991). To assess ADCC activity of a molecule of interest, an in vitro ADCC assay can be performed as described in US Pat. No. 5,500,362 or 5,821,337 or US Pat. No. 6,737,056 (Presta). Effector cells useful for such assays include peripheral blood mononuclear cells (PBMC) and natural killer (NK) cells. Alternatively, or additionally, ADCC activity of the molecule of interest can be assessed in vivo, for example in an animal model disclosed in Clynes et al., (USA) 95: 652-656 (1998).

"Treating" or "treatment" or "mitigation" refers to a therapeutic treatment that renders the subject helpless (weak) if the desired pathological condition or disease is not treated or the condition does not inhibit recurrence. Subjects are successfully “treated” for B cell malignancies after administration of a therapeutic amount of CD40 binding antibody, when the subject exhibits an observable and / or measurable decrease or absence of one or more signs and symptoms of the particular disease. For example, a significant decrease in the number of cancer cells or the absence of cancer cells; Reduction in tumor size; Inhibit (ie, slow to some extent and preferably stop) tumor metastasis; Some degree of inhibition of tumor growth; An increase in the duration of remission, and / or some relief, of one or more symptoms associated with the specific cancer; Reductions in morbidity and mortality, and improved quality of life. In addition, the reduction of the signs or symptoms may be felt by the patient. The treatment may achieve a complete response, defined as the disappearance of all signs of cancer, or a partial response in which the size of the tumor is preferably reduced by more than 50%, more preferably by 75%. The patient is also considered treated if the patient experiences a stable disease. In one criterion, the antibodies of the invention achieve greater than 95% peripheral blood B cell depletion and B cell recovery up to 25% of baseline. In some embodiments, treatment with an anti-CD40 antibody will not progress cancer in cancer patients after 3 months, preferably after 6 months, more preferably after 1 year, even more preferably after 2 years of treatment. It is effective enough. Such parameters for assessing successful treatment and amelioration of the disease can be readily determined by routine procedures well known to those skilled in the art.

As used herein, the term “non-Hodgkin's lymphoma” or “NHL” refers to a cancer of the lymphatic system other than Hodgkin's lymphoma. Hodgkin's lymphoma can generally be distinguished from non-Hodgkin's lymphoma by the fact that Reed-Sternberg cells are present in Hodgkin's lymphoma and the cells are not present in non-Hodgkin's lymphoma.

"Effective amount" means an amount effective at and for a period of time necessary to achieve the desired therapeutic or prophylactic result. The "therapeutically effective amount" of a therapeutic agent may vary depending on factors such as the disease state, age, sex and weight of the subject, and the ability of the antibody to elicit the desired response in the subject. A therapeutically effective amount is also an amount in which a therapeutically beneficial effect is greater than any toxic or detrimental effect of the therapeutic agent. A “prophylactically effective amount” means an amount effective at and for a period of time necessary to achieve the desired prophylactic result. Generally, but not necessarily, since a prophylactic dose is used in subjects prior to or at an earlier stage of disease, the prophylactically effective amount will be less than the therapeutically effective amount.

The term "housekeeping gene" refers to a group of genes whose proteins encode a protein whose activity is essential for the maintenance of cellular function. These genes are generally expressed similarly in all cell types.

"Correlate" or "correlated" means comparing the performance and / or results of the first analysis or protocol with the performance and / or results of the second analysis or protocol in any way. For example, the results of the first analysis or protocol may be used when performing the second protocol and / or the results of the first analysis or protocol may be used to determine whether the second analysis or protocol should be performed. For embodiments of gene expression analysis or protocol, the results of gene expression analysis or protocol can be used to determine whether a specific treatment regimen should be performed.

As used herein, the word "label" refers to a compound or composition that is conjugated or fused directly or indirectly to a reagent, eg, a nucleic acid probe or antibody, to facilitate detection of the conjugated or fused reagent. The label may be detectable by itself (eg, a radioisotope label or a fluorescent label) or may catalyze a detectable chemical alteration of the substrate compound or composition in the case of an enzyme label.

As used herein, the articles "a", "an", and "the" may mean singular or plural unless otherwise indicated (ie, may mean one or more).

C. Method of the Invention

The present invention provides methods for assessing or assisting in the responsiveness of a subject with B-cell lymphoma to treatment with an anti-CD40 antibody. The invention also provides methods for predicting responsiveness to anti-CD40 antibody treatment or monitoring treatment / response in subjects with B-cell lymphoma. The present invention provides a method of selecting a subject with B-cell lymphoma that is suitable for treatment with an anti-CD40 antibody and for follow-up treatment using anti-CD40 antibody treatment. In some embodiments, the method measures the expression level of one or more marker genes of any of Tables 2-4, 6, 7, and 13 in a sample comprising B lymphoma cells obtained from a subject; Predicting, evaluating, or assisting in evaluating a subject's responsiveness to anti-CD40 antibody treatment based on a measure of the expression level of said one or more marker genes. In some embodiments, the method comprises measuring the measured expression level of at least one marker gene of any of Tables 2-4, 6, 7, and 13 in a B-cell lymphoma sample from a subject to a reference level for each marker gene. Includes comparison

The method of the present invention assists the clinician in selecting patients during the development of anti-CD40 antibody therapy to identify patients with B-cell lymphoma for treatment with anti-CD40 antibodies, wherein individual patients are treated with specific treatment regimens. It is useful for predicting the likelihood of success in treatment, for assessing and monitoring disease progression, for monitoring efficacy of treatment, and for determining the prognosis of an individual patient. Any of these embodiments are included in the present invention.

In some embodiments, the B-cell lymphoma is non-Hodgkin's lymphoma (NHL), including but not limited to follicular lymphoma, recurrent follicular lymphoma, small lymphocytic lymphoma, mantle cell lymphoma, marginal lymphoma, lymphoid cell Lymphomas, mycelial sarcoma / Segri syndrome, splenic marginal lymphomas, and diffuse large cell B-cell lymphomas.

In some embodiments, B-cell lymphoma is painless. In some embodiments, B-cell lymphoma is aggressive. In some embodiments, B-cell lymphoma is highly aggressive. In some embodiments, the painless B-cell lymphoma is follicular lymphoma, marginal lymphoma, or small lymphocytic lymphoma. In some embodiments, the painless B-cell lymphoma is follicular lymphoma.

Marker  gene

The expression level of one or more marker genes in a B-cell lymphoma sample compared to the reference level may be used to predict, assess, or assist in evaluating the responsiveness of B-cell lymphoma to treatment with an anti-CD40 antibody. Used in

Genes differentially expressed (statistically significantly increased or decreased) in anti-CD40 antibody sensitive NHL cell lines compared to resistant NHL cell lines are shown in Tables 2-4, 6 and 7. "Anti-CD40 antibody susceptible cells" are cells with an IC25 value of less than 0.4 μg / ml in the reduction of cell viability by the anti-CD40 antibody tested as described in Example 1. “Anti-CD40 resistant cells” are cells in which the IC25 value exceeds 1 μg / ml in a decrease in cell viability when tested as in Example 1. Some of the genes of Tables 2-4, 6 and 7 are in the CD40 ligand downregulated pathway (eg, VNN2, MEF2C, LTB, KCNN3, NCF1, BCL6, IGJ, ELTI1902, PNOC, CSF2RB, and POU2AF1); Some of the genes in the table are in the B-cell receptor signal transduction pathways (eg, CD22, RGS13, and MEF2B). In addition, the association of expression levels of IFITM1, CD40, RGS13, VNN2, LMO2, CD79B, CD22, BTG2, IGF1R, CD44, CTSC, EPDR1, UAP1 and PUS7 (Table 13) was confirmed by the clinical trial described in Example 2 It became. Expression levels of one or more of these genes are used in the methods of the invention. In some embodiments, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 20, Expression levels of at least 25, or at least 30 genes are used in the methods of the invention.

In some embodiments, the expression level of one or more genes selected from the group consisting of VNN2, MEF2C, LTB, KCNN3, NCF1, BCL6, IGJ, ELTI1902, PNOC, CSF2RB, POU2AF1, CD22, RGS13, and MEF2B is measured and / or Used. In some embodiments, the expression level of one or more genes selected from the group consisting of IFITM1, CD40, RGS13, VNN2, LMO2, CD79B, CD22, BTG2, IGF1R, CD44, CTSC, EPDR1, UAP1, and PUS7 is measured and / or Used. In some embodiments, the expression level of at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, or 14 of these genes is Measured and / or used. In some embodiments, expression levels of CD22, CD40, and BCL6 are measured and / or used. In some embodiments, expression levels of CD40, RGS13, CD22, BTG2, IGF1R, and CD44 are measured and / or used. In some embodiments, expression levels of IFITM1, CD40, RGS13, VNN2, LMO2, CD79B, CD22, BTG2, IGF1R, CD44, CTSC, EPDR1, UAP1, and PUS7 are measured and / or used. In some embodiments, at least 1, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least of the genes of Table 7 or Table 13. 13, at least 14, or 15 expression levels are measured and / or used.

The genes (including sequences) identified in Tables 2-4, 6, 7, and 13 are known in the art. For example, an example of a Genbank accession number for a human gene is as follows:

The nucleic acid sequences of some genes mentioned in Tables 2-4, 6, 7 and 13 are shown in Figure 6 (6a-6ii).

Reference level

Measured expression levels of one or more marker genes in B-cell lymphoma samples are compared to reference levels. In some embodiments, the reference level is between types of B-cell lymphoma whose expression levels differ, eg, between B-cell lymphoma susceptible to anti-CD40 antibodies and B-cell lymphoma resistant to anti-CD40 antibodies. The level of expression of a gene that does not change (significantly unchanged). In some embodiments, the expression levels of one or more housekeeping genes shown in Table 8 are used as reference levels. In some embodiments, the expression levels of one or more housekeeping genes shown in Table 9 are used as reference levels.

In some embodiments, the measured expression level of the marker gene is normalized using the reference level. In some embodiments, the standardized expression level of the marker gene is calculated as the ratio or difference between the marker gene and the reference expression level, either original or logarithmic scale, respectively.

The reference genes of Tables 8 and 9 were selected as specific standardization counterparts for the marker genes of Table 4. Reference genes were selected for high mean expression and low fluctuations in B cell lymphoma samples. In addition, reference genes were chosen to have similar variations between replicate expression measurements of individual cell lines compared to variations between expression measures of biologically distinct cell lines. In addition, the reference gene was chosen to have a low statistical association with one or more of the markers in Table 4.

In some embodiments, the reference level is the measured expression level of the marker gene in different B-cell lymphoma samples. In some embodiments, different B cell lymphoma samples comprise B lymphoma cells that are resistant to anti-CD40 antibody induced cell death.

In some embodiments, the reference level is the level of expression of the corresponding marker gene in a sample comprising B lymphoma cells from a subject having increased tumor volume after anti-CD40 antibody treatment and / or reduced tumor volume after anti-CD40 antibody treatment. Determined on the basis of In some embodiments, the sample from the subject for reference level determination comprises B lymphoma cells of the same type as the sample from the subject whose responsiveness to anti-CD40 antibody treatment is predicted or evaluated. In some embodiments, the same methods (eg, qRT-PCR) and / or reagents (eg, primers) can be used to determine the expression level of a marker gene in a sample and to determine the expression level of the corresponding marker gene in a reference sample. And probes) are used.

Measurement of expression levels

The methods disclosed herein provide methods for examining the expression level of one or more of these marker genes in lymphoma samples (eg, B-cell lymphoma samples) relative to a reference level. Methods and analysis include examining the expression of marker genes, eg, one or more genes listed in any of Tables 2-4, 6, 7, and 13. Expression levels can be measured at the mRNA level and / or protein level.

The present invention provides a method for measuring expression levels from a mammalian tissue or cell sample (eg, cells and / or tissues associated with B-cell lymphoma). For example, to obtain patient samples, H & E staining is performed and used as a guide for tissue macrodissection to concentrate tumor content. Samples may be obtained for a variety of procedures known in the art, including but not limited to surgical resection, aspiration or biopsy. The sample can be fresh or frozen. In some embodiments, the sample is fixed and embedded in paraffin or the like. In a method, a mammalian tissue or cell sample is obtained and tested for expression of one or more biomarkers. The method can be performed in a variety of assay formats, including assays to detect mRNA expression, enzymatic assays to detect the presence of enzyme activity, and immunohistochemical assays. Determination of the expression of such biomarkers in the tissue or cell will be predictive such that the tissue or cell will be sensitive / responsive to treatment with an anti-CD40 antibody.

As discussed below, expression of various biomarkers in a sample can be analyzed by a number of methods, microarray (gene and / or tissue array analysis), in situ ( in situ ) hybridization, Northern analysis, PCR analysis of mRNA, immunohistochemical and / or Western analysis, quantitative blood based analysis (e.g. serum ELISA) (e.g. to determine levels of protein expression And) and / or many such methods, including but not limited to biochemical enzymatic activity assays, are known in the art and are understood by those skilled in the art. Representative protocols for assessing the status of genes and gene products are described, for example, in Ausubel et al. eds., 1995, Current Protocols In Molecular Biology, Units 2 (Northern Blotting), 4 (Southern Blotting), 15 (Immunoblotting) and 18 (PCR Analysis) have. The following protocols for the detection of specific biomarkers, such as those listed in Tables 2-4, 6, 7, and 13 in the samples, are provided for purposes of illustration.

In some embodiments, the methods of the present invention add a protocol to test for the presence and / or expression of mRNA in a tissue or cell sample, for example, the genes listed in any of Tables 2-4, 6, 7, and 13 It includes. In some embodiments, the expression of various biomarkers in a sample can be analyzed by microarray techniques, which techniques can be used to determine mRNA, such as any of Tables 2-4, 6, 7 and 13, in tissue or cell samples. Examine or detect mRNA. By using nucleic acid microarrays, test and control mRNA samples from test and control tissue samples are reverse transcribed and labeled to generate cDNA probes. The probe is then hybridized to an array of nucleic acids immobilized on a solid support. The array is shaped so that the sequence and position of each member of the array can be known. For example, a selection of genes that are likely to be expressed in certain disease states can be arranged on a solid support. Hybridization with a particular array member of a labeled probe indicates that the sample from which the probe is derived expresses the gene. Differential gene expression analysis of disease tissue can provide useful information. Microarray technology uses nucleic acid hybridization techniques and computational techniques to assess mRNA expression profiles of thousands of genes in a single experiment (see, for example, WO 01/75166, published October 11, 2001; see also, for example, See, US Pat. No. 5,700,637, US Pat. No. 5,445,934 and US Pat. No. 5,807,522, Lockart, Nature Biotechnology, 14: 1675-1680 (1996); discussions on array construction can be found in Cheung, VG et al., Nature Genetics 21 (Suppl). ): 15-19 (1999). DNA microarrays are small arrays containing gene fragments that are synthesized directly on, or spotted on, a glass or other substrate. Thousands of genes are usually presented in a single array. Typical microarray experiments include the following steps: 1) preparation of fluorescently labeled targets from RNA isolated from samples, 2) hybridization of labeled targets to microarrays, 3) washing, staining and scanning of arrays, 4) Analysis of the Scanned Image, and 5) Generation of Gene Expression Profiles. Two main types of DNA microarrays are currently used: gene expression arrays containing oligonucleotide (usually 25-70mer) arrays and PCR products made from cDNA. In forming an array, oligonucleotides can be prefabricated and spotted on the surface or synthesized directly onto the surface (in situ).

The Affymetrix GeneChip® system is a commercially available microarray system comprising an array made by direct synthesis of oligonucleotides on a glass surface. Probe / Gene Arrays: Oligonucleotides (usually 25-mers) are synthesized directly onto glass wafers by a combination of semiconductor-based photolithography and solid state chemical synthesis techniques. Each array contains up to 400,000 different oligomers, and each oligomer is present in millions of copies. Since oligonucleotide probes are synthesized at known positions on the array, hybridization patterns and signal intensities can be interpreted in terms of gene identity and relative expression levels by Affymetrix Microarray Suite software. Each gene is presented on an array by a series of different oligonucleotide probes. Each probe pair consists of a complete match oligonucleotide and a mismatch oligonucleotide. A complete match probe has a sequence that is exactly complementary to a particular gene and thus measures the expression of the gene. Mismatch probes differ from complete match probes by a single base substitution at the central base position, thus interfering with the binding of the target gene transcript. This helps to determine the background and nonspecific hybridization that contributes to the measured signal for the complete match oligomer. The microarray suite software subtracts the hybridization intensity of the mismatch probe from the hybridization intensity of the complete match probe to determine the absolute or specific intensity value for each probe cell. Probes are selected based on current information from Genbank and other nucleotide deposit institutions. The sequence is thought to recognize a unique region of the 3 'end of the gene. GeneChip hybridization ovens ("rotary" ovens) are used to perform hybridization of up to 64 arrays at a time. Fluidics stations perform washing and staining of probe arrays. It is fully automated and includes four modules, where each module holds one probe array. Each module is controlled independently through the Microarray Suite software using preprogrammed Fluidic protocol. The scanner is a confocal laser fluorescence scanner that measures the fluorescence intensity emitted by labeled cRNA bound to a probe array. A computer workstation with microarray suite software controls the fluidic station and scanner. The microarray suite software can control up to eight fluidic stations using preprogrammed hybridization, wash, and stain protocols for the probe array. The software also uses an appropriate algorithm to obtain hybridization intensity data and convert it to the presence / absence call for each gene. Finally, the software detects changes in gene expression between experiments by comparative analysis and formats the results into a .txt file that can be used in other software programs for further data analysis.

In some embodiments, expression of various biomarkers in a sample can also be assessed by examining gene deletion or gene amplification. Gene deletion or amplification can be performed by any one of a wide variety of protocols known in the art, such as conventional Southern blotting, Northern blotting to quantify transcription of mRNA (Thomas, Proc. Natl. Acad. Sci. USA, 77 (5201-5205 (1980)), dot blotting (DNA analysis), or in situ hybridization using appropriately labeled probes (eg, FISH), cytogenetic methods or comparative genomes using appropriately labeled probes Can be measured by hybridization (CGH). As one example, these methods can be used to detect the deletion or amplification of the genes listed in any of Tables 2-4, 6, 7, and 13.

In some embodiments, the expression of various biomarkers in a sample can be characterized by hybridization assays using complementary DNA probes (eg, in situ hybridization using labeled riboprobes, Northern blots, and related techniques) and various nucleic acid amplification assays (eg, , RT-PCR using complementary primers, eg, primers specific for one or more genes listed in any of Tables 2-4, 6, 7, and 13, and other amplification detection methods, such as branched DNA, SISBA , TMA, etc.).

Tissue or cell samples from mammals can be conveniently analyzed for mRNA of the genes listed in any of Tables 2-4, 6, 7, and 13, for example using Northern, dot blot or PCR analysis. In some embodiments, expression of one or more biomarkers can be analyzed by RT-PCR. In some embodiments, the RT-PCR can be quantitative RT-PCR (qRT-PCR). In some embodiments, the RT-PCR is a real time RT-PCR. In some embodiments, the RT-PCR is quantitative real time RT-PCR. RT-PCR assays, such as quantitative PCR assays, are well known in the art. In an exemplary embodiment of the invention, a method of detecting mRNA in a biological sample comprises generating cDNA from the sample by reverse transcription using at least one primer; Amplifying the thus produced cDNA using polynucleotides as sense and antisense primers to amplify the cDNA therein; Detecting the presence of the desired amplified cDNA. In some embodiments, the real time RT-PCR can be quantitative RT-PCR. In some embodiments, real-time RT-PCR can be performed using TaqMan® chemistry (Applied Biosystems). In some embodiments, real-time RT-PCR can be performed using TaqMan® Chemistry (Applied Biosystems) and ABI Prism® 7700 Sequence Detection System (Applied Biosystems). Real-time RT-PCR indicates that Taq polymerase has a 5'-3 'exonuclease activity and a double-labeled fluorescent oligonucleotide problem arises where the fluorescent signal is only released upon cleavage based on the principle of fluorescence resonance energy transfer. Principles are combined (see, eg, Overbergh, L. et al., J Biomolecular Techniques 14 (1): 33-43 (2003)). In addition, the method may include one or more steps that allow for determining the level of mRNA of a mRNA, eg, the genes listed in any of Tables 2-4, 6, 7, and 13, in a biological sample (eg, mRNA By examining the level of the "housekeeping" gene simultaneously with the comparable control mRNA sequences of the actin family members and / or one or more genes listed in Table 8 or Table 9). Examples of primers and probes that can be used to perform qRT-PCR are provided in Table 10.

In some embodiments, expression of proteins encoded by genes listed in any of Tables 2-4, 6, 7, and 13 in a sample is investigated using immunohistochemistry and staining protocols. Immunohistochemical staining of tissue sections has proven to be a reliable method of assessing or detecting the presence of proteins in a sample. Immunohistochemistry (“IHC”) techniques utilize antibodies to probes and generally visualize cellular antigens in situ by color or fluorescent methods.

For sample preparation, tissue or cell samples from mammals (generally human patients) can be used. Examples of samples include, but are not limited to, tissue biopsies, blood, lung aspirates, sputum, lymphatic fluid. Samples can be obtained by a variety of procedures known in the art, including but not limited to surgical excision, aspiration or biopsy. The tissue may be fresh or frozen. In some embodiments, the sample is fixed and embedded in paraffin or the like.

Tissue samples can be fixed (ie, preserved) by conventional methods (see, eg, "Manual of Histological Staining Method of the Armed Forces Institute of Pathology," 3 ^rd edition (1960) Lee G. Luna, HT (ASCP) Editor, The Blakston Division McGraw-Hill Book Company, New York ;; The Armed Forces Institute of Pathology Advanced Laboratory Methods in Histology and Pathology (1994) Ulreka V. Mikel, Editor, Armed Forces Institute of Pathology, American Registry of Pathology, Washington, DC). Those skilled in the art will understand that the fixative may be selected depending on the purpose for which the sample is histologically stained or otherwise analyzed. One skilled in the art will also understand that the length of fixation is determined by the size of the tissue sample and the fixative used. As an example, neutral buffered formalin, Boin or paraformaldehyde can be used to immobilize the sample.

Generally, the sample is first immobilized, then dehydrated through an ascending series of alcohols, and infiltrated and embedded in paraffin or other fragmentation media to allow for sectioning of the tissue sample. Alternatively, sections obtained by sectioning tissue can be fixed. By way of example, tissue samples may be embedded in paraffin and processed by conventional methods (see, eg, "Manual of Histological Staining Method of the Armed Forces Institute of Pathology", supra). Examples of paraffins that can be used include, but are not limited to, paraplasts, broroids and tissue may. After embedding the tissue sample, the sample may be sectioned by microcutter or the like (see, eg, "Manual of Histological Staining Method of the Armed Forces Institute of Pathology", supra). As an example of such a process, the thickness of the sections can be from about 3 microns to about 5 microns. After sectioning, the sections can be attached to the slides by a plurality of standard methods. Examples of slide adhesives include, but are not limited to, silane, gelatin, poly-L-lysine, and the like. As an example, paraffin embedded sections may be attached to bipolar slides and / or slides coated with poly-L-lysine.

When paraffin is used as the embedding material, tissue sections are generally deparaffinized and rehydrated in water. Tissue sections can be deparaffinized by any of a number of standard methods. For example, xylene and a progressively descending series of alcohols can be used (see, eg, "Manual of Histological Staining Method of the Armed Forces Institute of Pathology", supra). Alternatively, commercially available deparaffinized inorganics such as Hemo-De7 (CMS, Houston, Texas, USA) can be used.

In some embodiments, after sample preparation, tissue sections can be analyzed using IHC. IHC can be performed in combination with additional techniques such as morphological staining and / or fluorescence in situ hybridization. Two common IHC methods are available, namely direct and indirect analysis. According to a first analysis, the binding to the target antigen of the antibody (eg, a protein or fragment thereof encoded by one or more genes listed in Tables 1-4, 6 and 7) is measured directly. This direct assay uses labeled reagents, such as fluorescent tags or enzyme-labeled primary antibodies, which can be visualized without further antibody interactions. In a typical indirect assay, the unconjugated primary antibody binds to the antigen, and then the labeled secondary antibody binds to the primary antibody. When the secondary antibody is conjugated to an enzyme label, a chromogenic or fluorescent substrate is added to provide visualization of the antigen. Signal amplification occurs because several secondary antibodies can react with different epitopes on the primary antibody.

Primary and / or secondary antibodies used in immunohistochemistry will generally be labeled with a detectable moiety. Many labels are available and they generally fall into the following categories:

(a) Radioisotopes, for example ³⁵ S, ¹⁴ C, ¹²⁵ I, ³ H and ¹³¹ I. Antibodies are described, for example, in Current Protocols in Immunology Volumes 1 and 2, Coligen et al. Ed. Wiley-Interscience, New York, New York, Pubs. (1991) can be labeled with radioisotopes using the techniques described, and radioactivity can be measured using scintillation coefficients.

(b) colloidal gold particles

(c) rare earth chelates (Europium chelates), Texas red, rhodamine, fluorescein, dansyl, lysamine, umbeliferon, phycoerythrin, phycocyanin or commercially available fluorophores such as SPECTRUM 0RANGE7 And fluorescent labels including, but not limited to, SPECTRUM GREEN7 and / or any one or more derivatives of the foregoing. Fluorescent labels can be conjugated to antibodies using, for example, the techniques disclosed in Current Protocols in Immunology, supra. Fluorescence can be quantified using a fluorometer.

(d) Various enzyme-substrate labels are available and some of them are shown in US Pat. No. 4,275,149. Enzymes generally catalyze the chemical alteration of a chromogenic substrate, which can be measured using a variety of techniques. For example, enzymes can catalyze color change in a substrate, which can be measured with a spectrophotometer. Alternatively, the enzyme can alter the fluorescence or chemiluminescence of the substrate. Techniques for quantifying the change in fluorescence have been described above. The chemiluminescent substrate is electrically excited by a chemical reaction and then emits measurable light (eg, using a chemiluminometer) or energizes the fluorescent recipient. Examples of enzyme labels include luciferase (eg firefly luciferase and bacterial luciferase; US Patent 4,737,456), luciferin, 2,3-dihydrophthalazinedione, malate dehydrogenase, urease, peroxidase, Horseradish peroxidase (HRPO), alkaline phosphatase, β-galactosidase, glucoamylase, lysozyme, saccharide oxidase (e.g. glucose oxidase, galactose oxidase, and glucose-6) Phosphate dehydrogenase), heterocyclic oxidases (eg uricase and xanthine oxidase), lactoperoxidases, microperoxidases and the like. Techniques for conjugating enzymes to antibodies are described in O'Sullivan et al., Methods for the Preparation of Enzyme-Antibody Conjugates for use in Enzyme Immunoassay, in Method in Enzym. (ed J. Langone & H. Van Vunakis), Academic press, New York, 73: 147-166 (1981).

Examples of enzyme-substrate combinations are for example

(i) horseradish peroxidase (HRPO) and hydrogen peroxidase as substrate, where hydrogen peroxidase is a dye precursor (e.g. orthophenylene diamine (OPD) or 3,3 ', 5,5 '-Tetramethyl benzidine hydrochloride (TMB)));

(ii) para-nitrophenyl phosphate as chromogenic substrate and alkaline phosphatase (AP); And

(iii) chromogenic substrates (eg p-nitrophenyl-β-D-galactosidase) or fluorescent substrates (eg 4-methylumbeliferyl-β-D-galactosidase) and β- D-galactosidase (β-D-Gal)

It includes.

Many other enzyme-substrate combinations are available to those skilled in the art. For a general review of these, see US Pat. Nos. 4,275,149 and 4,318,980. Sometimes the label is conjugated indirectly to the antibody. Those skilled in the art will be aware of various techniques for achieving this. For example, the antibody can be conjugated with biotin, and any of the four broad categories of labels can be conjugated with avidin, or vice versa. Biotin binds selectively to avidin and thus, the label can be conjugated to the antibody in this indirect manner. Alternatively, to achieve indirect conjugation of the label with the antibody, the antibody is conjugated with a small hapten and one of the different label types described above is conjugated with an anti-hapten antibody. Thus, indirect conjugation of the label with the antibody can be achieved.

In addition to the sample preparation procedures discussed above, further processing of tissue sections may be required before, during or after IHC. For example, epitope repair methods, such as heating of tissue samples in citrate buffer, can be performed (eg, Leong et al., Appl. Immunohistochem. 4 (3): 201 (1996)). Reference).

After any blocking step, the tissue sections are exposed to the primary antibody for a sufficient time under conditions suitable for the primary antibody to bind to the target protein antigen in the tissue sample. Conditions suitable for achieving this can be determined by routine experimentation. The extent to which the antibody binds to the sample is determined using any one of the detectable labels discussed above. Preferably, the label is an enzyme label (eg HRPO) that catalyzes the chemical alteration of a chromogenic substrate, such as a 3,3'-diaminobenzidine chromogen. Preferably, the enzyme label is conjugated to an antibody that specifically binds to the primary antibody (eg, the primary antibody is a rabbit polyclonal antibody and the secondary antibody is a goat anti-rabbit antibody).

In some embodiments, an antibody used in an IHC assay that detects expression of one or more biomarkers is one or more biomarkers of interest, eg, one or more encoded by genes listed in any of Tables 2-4, 6, and 7. An antibody produced to bind primarily to a protein. In some embodiments, the antibody is a monoclonal antibody. Antibodies are readily available in the art, including those available from various commercial sources, and can also be generated using conventional techniques known in the art.

The sample prepared in this way can be arrange | positioned and a cover can be covered. The slide evaluation can then be carried out, for example using a microscope, and the staining intensity criteria commonly used in the art can be used. As an example, the dyeing intensity criteria can be evaluated as follows.

<Table A>

In another method, the sample may be contacted with an antibody specific for the biomarker under conditions sufficient to form an antibody-biomarker complex and then detect the complex. The presence of biomarkers can be detected by a number of methods, such as Western blotting and ELISA methods for analyzing a wide variety of tissues and samples, including plasma and serum. A wide variety of immunoassay techniques using this assay format are available (see, eg, US Pat. Nos. 4,016,043, 4,424,279 and 4,018,653). The methods include traditional competitive binding assays as well as single and double site or “sandwich” assays in uncompetitive forms. The assay also includes direct binding of the labeled antibody to the target biomarker.

Sandwich analysis is the most useful and commonly used analysis. Many variations of sandwich analysis techniques exist, all of which are included in the present invention. In brief, in a typical forward assay, an unlabeled antibody is immobilized on a solid substrate and the sample being tested is contacted with bound molecules. After incubation for an appropriate time, a second antibody specific for the antigen, labeled with a reporter molecule capable of producing a detectable signal for a time sufficient to form an antibody-antigen complex, is added and the other of the antibody-antigen-labeled antibody Incubate for a time sufficient to form the complex. Any unreacted material is washed away and the signal generated by the reporter molecule is detected to determine the presence of the antigen. The results can be qualitative by simple observation of the visible signal, or can be quantified by comparison with a control sample containing a known amount of biomarker.

Modifications of the anterior assay include simultaneous assays in which both the sample and the labeled antibody are added simultaneously to the bound antibody. This technique is known to those skilled in the art, including any small variations that are readily apparent. In a typical anterior sandwich assay, the first antibody with specificity for the biomarker is covalently or passively bound to a solid surface. Solid surfaces are generally glass or polymers and the most commonly used polymers are cellulose, polyacrylamide, nylon, polystyrene, polyvinyl chloride or polypropylene. The solid support may be in the form of tubes, beads, discs of microplates or any other surface suitable for performing an immunoassay. Binding methods are known in the art and generally consist of crosslinking, covalent bonding or physical adsorption, and the polymer-antibody complexes are washed at the time of test sample preparation. An aliquot of the sample to be tested is then added to the solid phase complex and subjected to sufficient time (e.g., from room temperature to 40 ° C, for example 25 ° C to 32 ° C) for conditions of binding of any subunit present in the antibody Incubate for 2-40 minutes or overnight if more convenient). After the incubation time, the antibody subunit solid phase is washed and dried to incubate with a second antibody specific for a portion of the biomarker. The second antibody is linked to a reporter molecule that is used to indicate binding to the molecular marker of the second antibody.

In some embodiments, the method involves immobilizing a target biomarker in a sample and then exposing the immobilized target to a specific antibody that may or may not be labeled with a reporter molecule. Depending on the amount of target and the intensity of the reporter molecule signal, the bound target can be detected by direct labeling with the antibody. Alternatively, a second labeled antibody specific for the first antibody is exposed to the target-first antibody complex to form a ternary complex of the target-first antibody-second antibody. The complex is detected by the signal emitted by the reporter molecule. As used herein, the term "reporter molecule" refers to a molecule that provides a signal that is identifiable through analysis by its chemical nature, allowing detection of antigen-bound antibodies. The reporter molecules most commonly used in this type of analysis are enzymes, fluorophores or radionuclide containing molecules (ie radioisotopes) and chemiluminescent molecules.

For enzyme immunoassays, enzymes are typically conjugated to a second antibody by glutaraldehyde or periodate. However, as will be readily appreciated, there are a wide variety of different conjugation techniques readily available to those skilled in the art. Commonly used enzymes include horseradish peroxidase, glucose oxidase, galactosidase and alkaline phosphatase. Substrates used with specific enzymes are generally selected for the generation of detectable color changes upon hydrolysis of the corresponding enzyme. Examples of suitable enzymes include alkaline phosphatase and peroxidase. In addition, it is also possible to use a fluorescent substrate that produces a fluorescent product rather than the above-described chromogenic substrate. In all cases, the enzyme labeled antibody is added to the first antibody-molecular marker complex to bind and then excess reagents are removed by washing. Then a solution containing the appropriate substrate is added to the complex of antibody-antigen-antibody. The substrate will react with the enzyme linked to the second antibody to generate a qualitative visible signal, which can generally be further quantified spectrophotometrically to provide an indication of the amount of biomarker present in the sample. Alternatively, fluorescent compounds such as fluorescein and rhodamine can be chemically coupled to the antibody without altering the binding ability of the antibody. When activated by irradiation with light of a particular wavelength, a fluorochrome-labeled antibody absorbs light energy to induce the molecule into an excitable state and then emits light in a characteristic color that is visually detectable with an optical microscope. Release. As in the EIA, the fluorescently labeled antibody binds to the first antibody-molecular marker complex. After washing off the unbound reagent and exposing the remaining ternary complex to light of the appropriate wavelength, the fluorescence observed indicates the presence of the desired molecular marker. Immunofluorescence and EIA techniques are both well established in the art. However, other reporter molecules may also be used, such as radioisotopes, chemiluminescent or bioluminescent molecules.

In some embodiments, expression of a selected biomarker in a tissue or cell sample can be investigated by functional or activity based analysis. For example, if the biomarker is an enzyme, assays known in the art can be performed to determine or detect the presence of a given enzymatic activity of a tissue or cell sample.

In any of the above methods for assessing the expression level of one or more biomarkers, a sample comprising a target molecule can be obtained by methods well known in the art, which is suitable for the particular type and location of the disease of interest. Tissue biopsies are often used to obtain representative pieces of diseased tissue. Alternatively, the cells can be obtained indirectly in the form of tissues / fluids known or thought to contain the disease cells of interest. For example, samples of disease lesions can be obtained by excision, bronchoscopy, microneedle aspiration, bronchial smears, or from sputum, pleural effusion or blood. The gene or gene product may be detected from diseased tissue or from other body samples such as urine, sputum or serum. The same techniques as discussed above for the detection of target genes or gene products in disease samples can be applied to other body samples. By screening the body sample, a simple initial diagnosis of these diseases can be performed. In addition, the progress of therapy can be more easily monitored by testing the body sample for target genes or gene products.

Means for concentrating tissue preparations for diseased cells are known in the art. For example, tissue can be isolated from paraffin or frozen sections. The cells of interest can also be isolated from normal cells by flow cytometry or laser capture microdissection. These and other techniques for separating diseased cells from normal cells are well known in the art. When diseased tissue is heavily contaminated with normal cells, detection of signature gene expression profiles may be more difficult, but techniques for minimizing contamination and / or false positive / negative results are known, some of which are described below. For example, the sample can also be assessed for the presence of biomarkers (including mutations) or vice versa that are known to be associated with the disease cells of interest but not to the corresponding normal cells.

Diseases that plague mammals, such as B-cell lymphoma, after determining that the tissue or cell sample expresses one or more biomarkers, indicating that the tissue or cell sample will be sensitive to treatment with an anti-CD40 antibody It is contemplated that an effective amount of an anti-CD40 antibody may be administered to a mammal, eg, a human, to treat the disease. Diagnosis of various pathological conditions described herein in a mammal, eg, a human, can be performed by one of skill in the art.

Compare expression levels, anti- CD40  Responsiveness of B-Cell Lymphoma to Antibody Treatment

Predict, evaluate, or assist in his evaluation

The methods described herein include comparing the measured expression level and the reference level of the marker gene. The reference level is the measured expression level of the reference gene different from the marker gene in the different sample or the measured expression level of the same marker gene.

In some embodiments, the measured expression level of a marker gene in a B cell lymphoma sample from a subject is compared to the measured expression level of a reference gene in the sample. In some embodiments, the expression level of the reference gene is substantially unchanged between various types of B lymphoma cells, including anti-CD40 antibody sensitive cells and resistant cells (eg, the genes of Table 8 or Table 9). In some embodiments, the ratio of the measured expression level of the marker gene to the measured expression level of the reference is calculated, which ratio can be used to assess or assist in responsiveness of B cell lymphoma to anti-CD antibody treatment. have.

In some embodiments, the measured expression level of the marker gene in the B cell lymphoma sample from the subject is compared to the measured expression level of the marker gene in the reference sample. In some embodiments, the reference sample comprises B lymphoma cells that are not resistant or reactive to anti-CD40 antibodies. For example, the comparison is performed to determine the magnitude of the difference between the measured expression level of the marker gene in the sample and the reference sample (eg, the expression level of the marker gene in the sample from the subject and the reference sample). Compare multiples or percentages of the difference between). An increase or decrease in the expression of a marker gene in a sample from a subject compared to the expression of a marker gene in a reference sample comprising B lymphoma cells that is not resistant or reactive to an anti-CD40 antibody may result in a B-cell for treatment with an anti-CD40 antibody. Present or indicate responsiveness of lymphoma. See Table 4 for marker genes with increased and decreased expression in anti-CD40 antibody sensitive cells compared to resistant cells. For example VNN2, MEF2C, LTB, KCNN3, NCF1, BCL6, IGJ, ELTI1902, PNOC, CSF2RB, POU2AF1, CD22, RGS13 and MEF2B are generally overexpressed in anti-CD40 antibody sensitive cells compared to resistant cells. In some embodiments, the fold increase in the expression level of a sample from a subject can be at least about 1.5X, 1.75X, 2X, 3X, 4X, 5X, 6X, 7X, 8X, 9X, or 10X of the expression level of a reference sample. . In some embodiments, the reduced fold of the expression level of a sample from a subject may be less than about 0.01, 0.05, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, or 0.8 of the expression level of the reference sample.

In some embodiments, the expression level of one or more marker genes selected from the group consisting of IFITM1, CD40, RGS13, VNN2, LMO2, CD79B, CD22, BTG2, IGF1R, CD44, CTSC, EPDR1, UAP1 and PUS7 is compared to a reference level. do.

In some embodiments, an increase in the expression level of one or more of IFITM1, CD79B, IGF1R, CD44, CTSC, EPDR1 and PUS7 relative to the reference level indicates that the subject is less responsive to agonist anti-CD40 antibody treatment. In some embodiments, the reference level is a value or range determined based on the expression level of the corresponding marker gene in a sample comprising B lymphoma cells from a subject having increased tumor volume following agonist anti-CD40 antibody treatment.

In some embodiments, increased expression of one or more CD40, RGS13, VNN2, LMO2, CD22, BTG2 and UAP1 relative to the reference level indicates whether the subject will respond to agonist anti-CD40 antibody treatment. In some embodiments, the reference level is a value or range determined based on the expression level of the corresponding marker gene in a sample comprising B lymphoma cells from a subject with reduced tumor volume after agonist anti-CD40 antibody treatment.

In some embodiments, the expression level of BCL6 is measured and compared to a reference level. The expression level of BCL6 is used to predict, assess, or assist in evaluating a subject's responsiveness to anti-CD40 antibody treatment. As shown in Example 2, BCL6 expression tends to be lower in subjects with increased tumors after agonist anti-CD40 antibody treatment. In some embodiments, an increase in expression of BCL6 relative to a reference level determined by the expression level of BCL6 in a sample from a subject with reduced tumor volume following agonist anti-CD40 antibody treatment results in the subject responding to agonist anti-CD40 antibody treatment. It can indicate whether or not to do so.

In some embodiments, the expression levels of the marker genes in Table 7 are measured, and a sensitivity index calculated as the sum of the sign t-scores for log2-scale expression of gene pairs 1-8 in Table 7 is determined, wherein − A sensitivity index greater than 4 suggests or indicates that B-cell lymphoma is responsive to anti-CD40 antibody treatment. In some embodiments, the sensitivity index is greater than -3, greater than -2, greater than -1, or greater than zero. In some embodiments, the sensitivity index is -4 to 20. In some embodiments, the sensitivity index is 0-20.

In some embodiments, the expression level of one or more of IFITM1, CD40, RGS13, VNN2, LMO2, CD79B, CD22, BTG2, IGF1R, CD44, CTSC, EPDR1, UAP1 and PUS7 is measured and the sensitivity index is measured expression of the marker gene. Calculated based on the level. For example, the following equation can be used to calculate the Susceptibility Index (SI):

Wherein the expression levels of at least one marker gene having a positive correlation value and at least one marker gene having a negative correlation value are determined; Wherein (i) β _j is the coefficient value for each marker gene measured; (ii) p is the number of marker genes measured; (iii) χ _i is a normalized expression level, transformed for a sample from the subject for the expression level of each marker measured; (iv) μ _j and σ _j are the mean and standard deviation for each marker gene measured; Wherein β _j , μ _j and σ _j are determined from patient samples comprising B lymphoma cells from clinical trials. In some embodiments, a value above zero for the sensitivity index indicates whether the subject will respond to anti-CD40 antibody treatment, or a value below zero for the sensitivity index indicates whether the subject will respond less to anti-CD40 antibody treatment. Indicates. Example 2 details the method for analyzing and determining the parameters for the reference sample and the new sample. In some embodiments, expression levels of IFITM1, RGS13, CD79B, CD22, BTG2, CD44, EPDR1 and UAP1 are measured and used for calculating the sensitivity index. In some embodiments, the same number of positively correlated marker genes and negatively correlated marker genes are measured and used for calculating the sensitivity index.

Methods of determining the sensitivity index are known in the art. Zhou H. and Hastie T. (2005) Regularization and variable selection via the elastic net; J.R. Statist. Soc. B. 67 (2). pp. 301-320; Friedman J., Hastie T. and Tibshirani R. 2008. Regularization Paths for Generalized Linear Models via Coordinate Descent. Technical Report, Department of Statistics, Stanford University (World Wide Web-stat.stanford.edu/~hastie/Papers/glmnet.pdf) R package glmnet]; R Development Core Team (2008). R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. ISBN 3-900051-07-0, URL World Wide Web at R-project.org.

The comparison can be done in any convenient way that is appropriate for the type of measured and reference value for the genetic marker in question. The comparison process can be manual or automatic. In some embodiments, the measured expression level is a standardized value. For example, expression levels can be normalized based on the equations below the transformed, standardized assay values described in Example 2. As will be apparent to those skilled in the art, replication measures can be considered for the expression level of the marker gene and / or reference gene. In some embodiments, replication measurements are taken into account for the measured values. Replication measurements can be considered by using the mean or median of the measured values as the “measured value”. Statistical analyzes known in the art can be used to demonstrate the significance of the difference between the two compared values.

term- CD40 Antibody Treatment

Marker genes identified in the present invention can be used to predict, assess, or assist in the responsiveness of B-cell lymphoma to treatment with one or more anti-CD40 antibodies. The anti-CD40 antibody may be one or more agonist antibodies (ie, bind to and stimulate CD40). Stimulating antibodies are of a different kind, such as: (1) transmit stimulatory signals through CD40, but increase the interaction between CD40 and CD40L (see, eg, US Pat. No. 5,182,368; and PCT WO 96/18413). Antibodies derived from the described G28-5 and G28-5), which do not reduce the interaction between CD40 and CD40L (eg, antibodies HuCD40-M2 and HuCD40-M3 and humanized antibodies described in US Pat. No. 5,674,492); And (2) capable of delivering stimulation signals through CD40 and increasing the interaction between CD40 and CD40L, for example from S2C6 (Francisco et al., 2000, Cancer Res. 60: 3225-31) and S2C6 May be an induced antibody. Agonist antibodies are also described in US Pat. No. 7,288,251. The anti-CD40 antibody may be one or more antagonist antibodies (ie, bind to CD40 and inhibit the activity induced by CD40L). Examples of antagonist anti-CD40 antibodies include human antibody CHIR-12.12 described in US Patent Application Publication 2007/0110754, and anti-CD40 antibody described in WO 97/31025.

The methods of the present invention may further comprise administering an effective amount of an anti-CD40 antibody to a subject with B-cell lymphoma after the subject has been identified as a candidate for treatment based on the assays / methods described herein. One or more anti-CD40 antibodies can be administered. In some embodiments, the anti-CD40 antibody is administered with one or more of the following therapeutic agents: rituximab, gemzar and ICE. For example, the anti-CD40 antibody may be rituximab therapy; With rituximab + gemza; With rituximab + ICE (ifosfamide, carboplatin, etoposide) (R-ICE); Or rituximab plus chemotherapy.

As used herein, “together” administration includes simultaneous administration and / or different time of administration. Co-administration also includes administration as a co-formulation (ie, different drugs are in the same composition) or administration as separate compositions, administration at different dosage frequencies or intervals, and administration using the same route or different routes.

Anti-CD40 antibodies or functional fragments may be used for the treatment of non-reactive or inappropriate reactions to treatment with any one of the following drugs or for relapses of NHL patients following treatment with these drugs: Rituximab (Genen tech); Okrelizumab (Genentech, Inc.); Ibritumomab thiuxetane (Zevalin ™, Biogen Idec); Tocitumomab (Bexxar ™, GlaxoSmithKline); HuMAX-CD20 ™ (GenMab); IMMU-106 (humanized anti-CD20 a.k.a.hA20 or 90Y-hLL2, Immunmedics); AME-133 (Applied Molecular Evolution / Eli Lilly); Gentuzumab ozogamicin (Mylotarg ™, humanized anti-CD33 antibody, Wyeth / PDL); Alemtuzumab (Campath ™, anti-CD52 antibody, Schering Plough / Genzyme); Epratuzumab (IMMU-103 ™, humanized anti-CD22 antibody, immunomedics).

The following references describe standard medical procedures for measuring lymphoma and CLL, their diagnosis, treatment and therapeutic efficacy (Canellos GP, Lister, TA, Sklar JL: The Lymphomas. WBSaunders Company, Philadelphia, 1998); van Besien K and Cabanillas, F: Clinical Manifestations, Staging and Treatment of Non-Hodgkin's Lymphoma, Chap. 70, pp 1293-1338, in: Hematology, Basic Principles and Practice, 3rd ed.Hoffman et al. (editors). Churchill Livingstone, Philadelphia, 2000; and Rai, K and Patel, D: Chronic Lymphocytic Leukemia, Chap. 72, pp 1350-1362, in: Hematology, Basic Principles and Practice, 3rd ed. Hoffman et al. (Editors) Churchill Livingstone, Philadelphia, 2000].

Anti-CD40 antibodies for use in therapy include chimeric, humanized and human antibodies. Any agonist or antagonist antibody described herein or known in the art can be used in the treatment. For example, the humanized anti-CD40 antibodies described in WO 2006/128103 can be used for the treatment of anti-CD40 antibodies, and these antibodies and their amino acid sequences are incorporated herein by reference. In some embodiments, the anti-CD40 antibody for use in the treatments described herein binds to CD40 (eg, human CD40) expressed on B lymphoma cells and induces apoptosis of B lymphoma cells. Anti-CD40 antibodies may also be characterized by killing B lymphoma cells in vivo via immune effector function, such as ADCC, CDC and / or ADCP. In some embodiments, the anti-CD40 antibody binds to CD40 with a K _d value of about 1 × 10 ⁻⁸ or less or 1 × 10 ⁻⁹ or less. In some embodiments, the anti-CD40 antibody binds to and stimulates CD40 (ie, agonist antibody). In some embodiments, the anti-CD40 antibody increases binding of the CD40 ligand to CD40, eg, at least 45%, at least 50%, at least 60%, or at least 75%. Methods for determining the increase in binding of CD40 ligands to CD40 are disclosed in US Pat. No. 6,838,261, the disclosure of which is incorporated herein by reference. In some embodiments, the anti-CD40 is a humanized antibody derived from murine monoclonal antibody S2C6 described in WO 00/75348 (including the antibodies provided in Tables 3 and 4 of WO 00/75348). In some embodiments, the anti-CD40 antibody comprises the heavy chain amino acid sequence shown in SEQ ID NO: 1 and the light chain amino acid sequence shown in SEQ ID NO: 2, eg, anti-CD40 Ab.1.

D. Kit

For use in the applications described or suggested above, kits or articles of manufacture are provided herein. Such kits may comprise at least one reagent specific for detecting the expression level of a marker gene described herein and may further comprise instructions for carrying out the methods described herein.

In some embodiments, the invention selectively or specifically describes primers and primer pairs that allow specific amplification of a polynucleotide of the invention or any specific portion thereof, and nucleic acid molecules of the invention or any portion thereof. Provided are compositions and kits comprising a hybridizing probe. The probe may be labeled with a detectable marker such as radioisotopes, fluorescent compounds, bioluminescent compounds, chemiluminescent compounds, metal chelators or enzymes. Such probes and primers are used to detect the presence of polynucleotides in a sample, eg, polynucleotides corresponding to the genes listed in Tables 1-4, 6, 7, and 13, and Tables 1-4, 6, 7, and 13 It can be used as a means for detecting a cell expressing a protein encoded by a polynucleotide corresponding to the genes listed in. As will be appreciated by those skilled in the art, a large number of different primers and probes may be prepared based on the sequences provided herein and used effectively to amplify and / or clone mRNAs and / or to determine their presence and / or level. have.

In some embodiments, the kit comprises a reagent for detecting the expression level of at least 2, at least 3, at least 5, at least 10, at least 15, at least 20 marker genes. The kit can also include a reference sample that is useful as generating a reference value. Marker genes are VNN2, MEF2C, LTB, KCNN3, NCF1, BCL6, IGJ, ELTI1902, PNOC, CSF2RB, POU2AF1, CD22, RGS13, MEF2B, LRRC8A, CD40, IFITM1, SMN1, PRRCA, EPDR1, PRPSR2, IGF2GF , YIPF3, CD79B, CD44, CTSC, UAP1 and PUS7. Reagents for detecting mRNA expression levels of a marker gene may comprise at least one pair of primers specific for amplifying the mRNA product of one marker gene. In some embodiments, pairs of primers can target the 3 ′ end of the mRNA sequence (eg, target mRNA in a 3 ′ UTR that is commonly shared with all transcript variants). In some embodiments, the kit may further comprise a surface or substrate (eg, microarray) to capture the probe for detection of the amplified nucleic acid.

In some embodiments, the kit comprises at least one pair of primers and probes specific for detecting the expression level of one marker gene using qRT-PCR. Examples of sets of primers and probes that can be used in qRT-PCR are shown in Table 10. To detect IFITM1, the primer and probe sets set forth in SEQ ID NOs: 27, 28 and 29, SEQ ID NOs: 60, 61 and 62, and SEQ ID NOs: 93, 94 and 95 can be used. To detect CD40, a set of primers and probes set forth in SEQ ID NOs: 24, 25 and 26, SEQ ID NOs: 57, 58 and 59, SEQ ID NOs: 90, 91 and 92 can be used. To detect RGS13, a set of primers and probes set forth in SEQ ID NOs: 114, 115, and 116, and SEQ ID NOs: 126, 127, and 128 can be used. To detect VNN2, a set of primers and probes set forth in SEQ ID NOs: 30, 31 and 32, SEQ ID NOs: 63, 64 and 65, and SEQ ID NOs: 96, 97 and 98 can be used. To detect LMO2, a set of primers and probes set forth in SEQ ID NOs: 12, 13 and 14, SEQ ID NOs: 45, 46 and 47, and SEQ ID NOs: 78, 79 and 80 can be used. To detect CD79B, a set of primers and probes set forth in SEQ ID NOs: 141, 142 and 143, SEQ ID NOs: 150, 151 and 152, and SEQ ID NOs: 159, 160 and 161 can be used. To detect CD22, the primer and probe sets set forth in SEQ ID NO: 15, 16 and 17, SEQ ID NOs: 48, 49 and 50, and SEQ ID NOs: 81, 82 and 83 can be used. To detect BTG2, the primer and probe sets set forth in SEQ ID NOs: 9, 10 and 11, SEQ ID NOs: 42, 43 and 44, and SEQ ID NOs: 75, 76 and 77 can be used. To detect IGF1R, a set of primers and probes set forth in SEQ ID NOs: 6, 7 and 8, SEQ ID NOs: 39, 40 and 41, and SEQ ID NOs: 72, 73 and 74 can be used. To detect CD44, a set of primers and probes set forth in SEQ ID NOs: 174, 175 and 176, SEQ ID NOs: 180, 181 and 182, and SEQ ID NOs: 186, 187 and 188 can be used. To detect CTSCs, the primer and probe sets set forth in SEQ ID NOs: 165, 166 and 167, SEQ ID NOs: 168, 169 and 170, and SEQ ID NOs: 171, 172 and 173 can be used. To detect EPDR1, the primers set forth in SEQ ID NOs: 21, 22 and 23, SEQ ID NOs: 54, 55 and 56, SEQ ID NOs: 87, 88 and 89, SEQ ID NOs: 129, 130 and 131, SEQ ID NOs: 132, 133 and 134, SEQ ID NOs: 135, 136 and 137 And probe sets can be used. To detect UAP1, the primer and probe sets set forth in SEQ ID NOs: 138, 139 and 140, SEQ ID NOs: 147, 148 and 149, and SEQ ID NOs: 156, 157 and 158 can be used. To detect PUS7, a set of primers and probes set forth in SEQ ID NOs: 177, 178 and 179, SEQ ID NOs: 183, 184 and 185, and SEQ ID NOs: 189, 190 and 191 can be used. To detect BCL6, the primer and probe sets set forth in SEQ ID NOs: 102, 103 and 104, and SEQ ID NOs: 108, 109 and 110 can be used.

Reagents for detecting the protein expression level of a marker gene can include an antibody that specifically binds to a protein encoded by the marker gene.

The kit may further comprise a conveying means which is compartmentalized to receive one or more container means, for example vials, tubes, etc., each said container means comprising one of the distinct elements to be used in the method. do. For example, one of the container means can comprise a probe that can be detectably labeled or labeled. Such probes may be antibodies or polynucleotides specific for the marker gene. If the kit uses nucleic acid hybridization to detect a target nucleic acid, the kit may also contain a container containing nucleotide (s) for amplification of the target nucleic acid sequence, and / or a reporter molecule, eg, an enzyme, fluorescence, or radioisotope. It may have a container comprising reporter-means, for example biotin-binding proteins, such as avidin or streptavidin, bound to an elemental label.

Kits of the present invention will typically include one or more other containers including the above-described containers and commercially and user-friendly materials, including buffers, diluents, filters, needles, syringes, and package inserts with instructions for use. will be. A label indicating that the composition is to be used for a particular therapeutic or non-therapeutic use may be present on the container and may also indicate instructions for in vivo or in vitro use as described above.

The kit may further comprise a set of instructions and materials for preparing a tissue or cell sample and for preparing nucleic acid (eg, mRNA) from the sample.

The present invention provides a variety of compositions suitable for use in carrying out the methods of the present invention, which can be used in kits. For example, the present invention provides a surface, such as an array, that can be used in such a method. In some embodiments, the array of the invention comprises individual nucleic acid molecules or collections of nucleic acid molecules useful for detecting mutations of the invention. For example, an array of the invention may comprise a series of discretely arranged individual nucleic acid oligonucleotides or a set of nucleic acid oligonucleotides that are hybridizable to a sample comprising a target nucleic acid, wherein said hybridization is in the presence of a mutation of the invention. Or absent.

Several techniques for attaching nucleic acids to solid substrates, such as glass slides, are well known in the art. One method is to incorporate modified bases or analogs containing moieties that can be attached to a solid substrate, such as amine groups, derivatives of amine groups, or other groups with positive charges, into the nucleic acid molecules to be synthesized. The synthesized product is then contacted with a solid substrate, such as a glass slide, coated with an aldehyde or other reactor that forms a covalent connection with the reactor on the amplification product and covalently attaches to the glass slide. Other methods are known in the art, such as using amino propyl silicon surface chemistry, as disclosed on the Internet web sites cmt.corning.com and cmgm.stanford.edu/pbrown1.

In addition, attachment of the group to oligonucleotides which can be later converted to the reactor can also be carried out using methods known in the art. Any attachment of the oligonucleotide to the nucleotide will be part of the oligonucleotide, which can then be attached to the solid surface of the microarray. The amplified nucleic acid may be further modified, for example, by cleavage into fragments or by attachment of a detectable label before or after attachment to a solid substrate, as necessary and / or acceptable in the technique used.

The following are examples of the methods and compositions of the present invention. It is understood that various other embodiments may be practiced, given the general description provided above.

Example

Example  1. CD40  For antibody treatment NHL  Predictive Genes for Patient Responsiveness Marker Ok

Substances and Methods

Cell Viability Assay

NHL cells are seeded in 50 μl RPMI 1640 supplemented with 2% FBS in 384 well plates at 1500-5000 cells / well and with a series of concentrations of crosslinked anti-CD40 Ab.1 or control antibody (anti-gD 5B6) Treated. For crosslinking, goat anti-human IgG Fcγ fragment-specific antibody (Jackson Immunoresearch (1) in a ratio of 1: 4 in medium at room temperature for 30 minutes prior to addition of anti-CD40 Ab.1 or anti-gD to cells. Incubated with F (ab ') 2 fragment of Jackson ImmunoResearch, West Grove, Pennsylvania). After 96 hours of incubation, cell viability was assessed using CellTiter-Glo Luminescent Cell Viability Assay (Promega, Madison, WI) according to the manufacturer's instructions. Each data point was performed in triplicate.

XLfit was used to calculate IC50, IC25 and maximum inhibition. Data is expressed as the mean of three independent experiments. Sensitivity to anti-CD40 Ab.1 was classified into three categories: sensitivity, intermediate, and resistance based on IC25 and IC50 values.

Antibodies

Anti-CD40 Ab.1 is a humanized IgG1 mAb for CD40. It is produced and secreted in Chinese hamster ovary (CHO) cell lines processed by genetic engineering. Anti-CD40 Ab.1, used in the Examples and referred to as anti-CD40 Ab.1, has the following amino acid sequence:

Heavy chain (SEQ ID NO: 1). The underlined ASN 294 residue in italics indicates the position of the carbohydrate moiety.

Light chain (SEQ ID NO: 2).

Generation and Analysis of Gene Expression Profiles

Total RNA was extracted with mir Vana ™ miRNA isolation kit (Ambion, Austin, TX) and analyzed using Affymetrix HGU133P2 whole genome expression microarray. Raw data was extracted using an Affymetrix scanner and the resulting CEL files were processed using gcRMA at the default of the R Bioconductor Package (Internet web site bioconductor.org). Significantly differentially expressed genes were identified using mediated t-tests for differences across anti-CD40 Ab.1 susceptibility and viability classes. Additional parameters were evaluated using the LIMA package and t-statistics, p-values, adjusted p-values, and B-statistics were calculated for each gene. Probes were mapped for each gene and 1: 1 probe-to-gene mapping was selected for downstream analysis using the most strongly associated probe while measuring sensitivity. For classification into sensitive or intermediate versus resistant groups, quantitative stepwise linear modeling was combined with qualitative analysis of the target pathway to identify a parsimonious set of genes to include in the analysis. Further details and results are given in the Results section below (Table 7).

Gene set enrichment analysis was determined using the GSEA module of Gene Pattern (www.genepattern.org). Enrichment scores confer prespecified classes of genes when their members are significantly differentially expressed in a consistent manner in the phenotype. The standardized enrichment score is calculated by adjusting the enrichment score to the number of genes in the gene set. The nominal p-value is determined by recalculating the normalized enrichment scores to replace the susceptibility and resistance markers and produce a null distribution.

Stepwise linear Modeling  Anti-identified using CD40Ab .1 sensitivity index

Each target gene is presented with its corresponding inversely correlated (semi-correlated) pair of genes in the order in which the target gene is selected for inclusion in the index (Table 7). The first three major genes (VNN2, RGS13, CD22 in Table 7) were selected from Tables 2-4 (Step 1) to model the predominant components of differential over-expression in sensitive and intermediate cell lines. The expression of these three genes is highly correlated with a correlation coefficient of at least +0.77. Due to their similarity, single pair gene EPDR1 was selected from Tables 2-4 to measure contrast overexpression in resistant cell lines. Including such anti-correlated pair genes in the assay provides auto-standardization in which both susceptibility and resistance are associated with high expression of one arm of the pair. By this mechanism, the assays do not rely on low total mRNA analysis levels to define any class, but instead describe each by a pattern of relative expression of major genes for their anti-correlation pairs (ie Sum of the sign t-scores of the _two logarithmic scales, where the sign corresponds to a multiple change estimate). In steps 2-5, additional pairs of genes were selected based on the mechanism of action from a new list of those with significant association to IC25 after adjustment for the cumulative sum of the sign t-scores for the genes identified in the previous step. . The stepwise procedure requires each new pair of genes to add additional predictive power to the sensitivity index. After step 5, no further gene pairs are needed for IC25 prediction. In step 6, a single additional pair was added for its ability to predict cell viability at maximum inhibition after adjusting for cumulative indices based on the preceding seven pairs of genes. BCL6 was added as a single pair without corresponding pairs on the basis of mechanism of action: it is currently not included in the final sensitivity index given as the sum of the sign t-scores for log2-scale expression of gene pairs 1-8. This may be explicitly included as an index based on clinical experience. For classification into sensitive or intermediate versus resistant groups, a preliminary cutoff for the sensitivity index was chosen to maximize the overall accurate sorting rate. Alternative classification rules based on selected probes can be optimized later for clinical application.

Results and analysis

In order to understand the mechanism of action of anti-CD40 antibodies and to identify one or more predictive markers for NHL patients' responsiveness to anti-CD40 antibody therapy, the inventors of anti-CD40 Ab.1 have been identified throughout a panel of 31 NHL cell lines. Activity was tested and cell viability in response to titration of anti-CD40 antibody was assessed. The IC25 values highlighted in Table 1 from this experiment indicated that the anti-CD40 antibody sensitized 10 cell lines with decreasing cell viability at concentrations <0.4 μg / ml (herein defined as 'sensitive' cell lines), and 13 cell lines Even cell concentrations of 1 μg / ml were not reduced (herein defined as 'resistant' cell lines). IC25 of eight cell lines are> 0.4 to <0.8 and will be defined herein as 'medium' cell lines.

Table 1 provides anti-CD40 Ab.1 IC25 susceptibility data across NHL cell lines in vitro. Specific lymphoma subtypes, IC25 values, and classifier data for each cell line are given for each cell line. DLBCL (subtypic giant B-cell lymphoma), FL (follicular lymphoma), MCL (mantle cell lymphoma), ALCL (large cell degenerative lymphoma).

To identify genes that predict anti-CD40 Ab.1 activity in vitro, RNA was prepared from cell lines in the cell division log phase and gene expression profiled using an Affymetrix HGU133P2 microarray. Genes differentially expressed between susceptible and resistant cell lines were determined by mediated t-test, and significance was determined using an adjusted P-value cutoff of ≦ 0.05 (Table 2). In Table 2, the gene list was filtered to adjusted p-value <0.05 (5% FDR), resulting in 110 unique genes. Probe ID, genetic symbol and description are presented. In addition, significant genes correlated with IC25 values across all NHL cell lines were determined by Spearman rank correlations and the genes were filtered using rho values of ≦ -0.57 or ≧ 0.57 (Table 3). In Table 3, the gene list was filtered using rho values of ≦ -0.57 or ≧ 0.57 to generate 130 unique genes. Probe ID, genetic symbol and description are also presented. Table 4 shows the combination of unique genes identified by each or both methods. In Table 4, Log (2) fold changes are shown, where positive fold changes indicate increased expression in the NHL cell line's susceptibility class with respect to anti-CD40 Ab.1 susceptibility, and negative fold changes increase in the resistance class Expression is shown. Genes represent 195 unique genes. Probe ID, genetic symbols and descriptions are also presented.

The highly expressed genes in Table 4 may be co-regulated genes that may not be involved in the biology of anti-CD40 activity. Thus, to understand the biological function of genes differentially expressed between susceptible and resistant cells, we performed gene set enrichment analysis (GSEA). In this analysis, we addressed the problem by calculating the mean t-statistics for the genes in the set and then comparing the mean t-statistics with the mean statistics calculated for a random set of genes of the same size. Low p-values may indicate that some correlation exists between the set of genes used to generate the statistics and the sample classification. Thus, gene set analysis can be interpreted as a summary of the properties of highly differentially expressed genes. Table 5 provides a gene set enrichment analysis of anti-CD40 Ab.1 sensitive versus resistant NHL cell lines. Enriched gene sets, number of genes per gene set, standardized enrichment score (NES), and nominal p-value (NOM p-val) are shown. The higher the NES and the lower the NOM p-val, the more likely the finding is significant.

Among the biologically relevant GSEA identified gene sets, gene sets involved in B-cell receptor signal transduction (BCR) and genes of embryonic center origin (Table 5) were concentrated. Of primary interest is the observation of genes involved in CD40 signal transduction as determined by the BASSO_GERMINAL_CENTER_CD40_DN gene set (FIG. 1) (Basso et al., Blood 104: 4088-96, 2004). This set of genes represents the genes reported to be inhibited by CD40L in Ramos cell lines. Ranking from the differentially expressed gene list and adjusted p-values are shown in Table 6 for this gene set. In Table 6, genes differentially expressed between sensitive and resistant cell lines were enriched for genes known to be CD40L downregulated. Ranked genes are derived from mediated t-tests (Table 2). A total of 70 genes are part of the gene set and the top 11 are shown in the table. The genes shown in Table 6 are overexpressed in anti-CD40 Ab.1 sensitive cell line. Partial overlap of the genes with the BCR and CD40L genes is expected because two signaling pathways converge at the axis of NF-κB transcription and both pathways can synergize to activate B-cells. Subsequently, it was confirmed whether any CD40L-derived gene could distinguish between a sensitive NHL cell line and an resistant NHL cell line for anti-CD40 Ab.1. Among the CD40L genes in the differentially expressed gene lists in Tables 2 and 3, VNN2 provided the most accurate distinction for sensitive and resistant cell lines (FIG. 2).

Further testing of the differentially expressed gene list also indicated that genes such as CD22, RGS13, and MEF2B (Table 2 and Figures 3, 4, 6) that expressed embryonic center B (GCB) cells were excessive in the anti-CD40 Ab.1 sensitive cell line. It was found to be expressed. CD40 signature genes were correlated with anti-CD40.Ab.1 susceptibility as shown in FIG. 5. In particular, RGS13 is one of the highest ranking genes by mediated t-test (Table 2), and the Spearman rank correlation (Table 3) throughout the cell line and as a single gene can distinguish between susceptibility and resistance and also moderate Type and tolerance classes can be distinguished with high accuracy, 96% accuracy for sensitivity versus tolerance, 81% accuracy for intermediate vs. tolerance, and 87% accuracy for sensitivity / medium vs. tolerance.

To obtain optimal classification accuracy, perhaps a genetic signature, or metagene, classifier will be required. Thus, in order to identify genes that may contribute to the most accurate classifier, algorithms were generated that identify pairs of genes that, when combined, will provide the best possible classification across cell lines with respect to anti-CD40 Ab.1 sensitivity. Therefore, the inventors performed stepwise linear modeling to achieve the above object, and the final gene selection is shown in Table 7. In Table 7, each target gene is shown with its corresponding inversely correlated (semi-correlated) pair genes in the order of steps in which the target gene is selected for inclusion in the index, as described above. The selection of these gene pairs is strongly potent in the sensitivity, intermediate and resistance classes for anti-CD40 Ab.1 when the sensitivity index, which is essentially the sum of the sign t-scores for log2-scale expression of gene pairs 1-8, is calculated Revealed the classification (Figure 4).

Taken together, CD40L plays an important role in activating B-cells and causes Ig class switching as well as swelling and proliferation of B-cells, and the CD40L signal transduction pathway also contains pro- and naïve and memory B-cells. And post-GCB-cells. Thus, NHL cells showing susceptibility to anti-CD40 Ab.1 are similar to native GCB-cells by gene expression profiling, and CD40L downregulated genes are highly expressed in contrast to resistant cells (anti-CD40 Ab It is surprising to know the relationship between the GCB and CD40 pathway activation states that determine susceptibility to .1).

To further identify predictive classifiers, xenograft models are used to explore the therapies (eg combination therapies). Real-time quantitative RT-PCR (qRT-PCR) is used to measure gene expression levels. After identifying predictive classifiers, immunohistochemistry (IHC) analyzes are developed for selected small groups of markers (eg, VNN2 and RGS13). Selected marker genes are further tested in clinical trial samples.

QRT-PCR and IHC are performed to determine the expression level of selected marker genes in clinical trial samples. The expression level in a sample from a patient with recurrent diffuse large cell B-cell lymphoma that is responsive to anti-CD40 treatment is compared to the expression level in a sample from a patient who is not responsive to the treatment.

Example  2. Anti-in clinical trials CD40 Ab .1 associated with responsiveness to treatment with hemp Kerr's confirmation

Clinical trial 001 ( II Prize)

A multicenter phase II open-label study to determine the total response rate and toxicology profile of anti-CD40 Ab.1 in patients with recurrent DLBCL. Tumor samples were evaluated in a central laboratory for pathological confirmation and CD40 expression. Eligible patients had a de novo or transformed DLBCL at diagnosis and were excluded if there was a history of painless lymphoma. The required prior therapy consisted of rituximab and, if eligible, combination chemotherapy with autologous stem cell transplantation. Patients received anti-CD40 Ab.1 intrapatient dose loading (1 mg / kg Day 1; 2 mg / kg Day 4; 4 mg / kg Day 8) and then 5 weeks at 8 mg / kg / wk IV was injected six times over (cycle 1). Responding patients and patients with SD (stable disease) were eligible for therapy that continued until disease progression or up to 12 cycles. Tumor tissue was taken from patients prior to treatment with anti-CD40 Ab.1. For example, samples were taken as part of routine lymphoma diagnosis.

Clinical Trial 002 (Phase I)

A multicenter multi-dose phase I study was conducted to test the safety, pharmacokinetics, immunogenicity and antitumor activity of intravenous anti-CD40 Ab.1 in patients with recurrent NHL. Multiple histological subtypes, including diffuse large cell B-cells (DLBCL; 14), follicular (FCL; 9), mantle cells (MCL; 9), marginal regions (MZL; 2), and lymphocytic (SLL; 1) Patients with type NHL were involved in this study. Patients were treated with a dose-loading schedule: 1, mg / kg of anti-CD40 Ab.1 on days 1 and 4, and 3, 4, 6 or 8 mg / kg over 4 cohorts Subsequent patient internal dose-increasing for 2-5 weeks at maximum dose. Subsequently, the rapid dose-loading schedule was tested in one cohort (40% increase in total anti-CD40 Ab.1 administered during Cycle 1). Responding patients or patients with stable disease were eligible for the second cycle, which consists of four consecutive weekly infusions at the cohort-specific maximum dose of anti-CD40 Ab.1. Eight DLBCL patients completed Cycle 1 and received a maximum dose of at least 3 mg / kg anti-CD40 Ab.1 with an objective response rate of 37.5% (ie, 1 CR and 2 PR) and 2 SD. Additional objective responses were seen in one MCL patient (CR) and one MZL patient (PR). The duration of median response for these five patients has not yet been reached (range 8-37 weeks). Tumor tissue was taken from patients prior to treatment with anti-Cd40 Ab.1. For example, samples were taken as part of routine lymphoma diagnosis.

Clinical sample preparation and qRT - PCR

Formalin fixed paraffin embedded (FFPE) recording tumor tissues from phase I and II clinical trials described above were obtained from the clinical investigation site with appropriate IRB approval and patient consent. 4-6 micron sections from tumor tissue were mounted on glass slides and one slide for each case was H & E stained using standard pathological experimental protocols. A board certified pathologist displays H & E slides for tumor content and performs RNA extraction using Ambion RecoverAll ™ Total Nucleic Acid Isolation Kit (Cat. No. AM1975; Applied Biosystems / Ambion) on FFPE tissue. The remaining tumor-containing zone was used as a guide for gross ablation.

450 ng of total RNA per sample was reverse transcribed using Applied Biosystems' High Capacity Reverse Transcription cDNA Synthesis Kit (Cat. No. 4368814; Applied Biosystems, Foster City, CA, USA) at a total reaction volume of 20 μl. Follow the manufacturer's recommendations, but perform a 60 min RT reaction shortened at 37 degrees. 5 ng of total RNA equivalent cDNA (assuming 100% cDNA synthesis efficiency) product was mixed with Applied Biosystems' 2X universal master mix (no UNG) in a volume of 15 μl for each PCR assay well. All amplifications were performed in triplicate in 384-well plates using a two-step (95 degrees 15 seconds, 60 degrees 1 minute) PCR amplification procedure. The reaction was performed at 40 cycles in a validated ABI 7900 real time PCR system. The sequences of the primers and probes used are shown in Table 10.

Data  process

As a result, the raw qRT-PCR was pre-treated according to the description under standardization, transformation and imposition, and the sensitivity index was calculated as described under the sensitivity index and classifier. Spearman rank correlations were used for correlation estimates and corresponding P-values. For multivariate susceptibility indices, probes were selected and coefficients were determined by Zhou et al., Statist. Soc. B. 67: 301-320, 2005, and Friedman, Hastie and Tibshirani, Regularization Paths for Generalized Linear Models via Coordinate Descent. Technical Report, Dept. of Statistics, Stanford University at www-stat.stanford.edu/~hastie/Papers/glmnet.pdf] for penalized regression of lasso (L1) and ridge (L2) penalties. Estimate using elastic net blend. The X ² test was used to test for associations between variable values of categories.

Standardize, Convert, and Replace

The following are the definitions of the analytical data and model parameters:

Justice

analysis Data

ℓ = reference set of samples (e.g., NHL cell line)

N _ℓ = sample size

p = number of probes (does not include normalizer)

N _lj ^{( Obs )} = detected sample size for probe j

N _ℓj ^{( ND )} = undetected sample size for probe j

y _ij ^{( Obs )} = raw analytical value detected for sample i, probe j

p _i ^{( nrm . Obs )} = number of detected normalizer values for sample i

y _ij ^{( nrm . Obs )} = detected standardized value for sample i, probe j

Model parameters

= 프로브 j에 대한 검출된 로그2 분석 값의 세트 ℓ 평균 (표준화되지 않은)

= Set ℓ average of detected log 2 analysis values for probe j (unstandardized)

= 프로브 j에 대한 검출된 로그2 분석 값의 세트 ℓ 표준 편차

= Set of detected log 2 analysis values for probe j l standard deviation

= 평균을 초과하는 표준 편차의 세트 ℓ 수

= Set ℓ number of standard deviations above the mean

For use in fitting reference sets of samples, e.g. exponential coefficients and classifier cutoffs, mean and standard deviation model parameters are computed using reference set data (see equation for reference set model parameters below). ). For a new sample, for example a single new sample for which the exponent and class are to be calculated, the model parameter should be taken from the reference set l and the set is chosen to be the most representative of the population from which the new sample is extracted. For example, the line of therapy for which the assay is used and the clinical reference set for each indication may be maintained. The equations for calculating the reference set model parameters and the transformed, standardized analysis values are presented below.

expression

Reference Set Model Parameters

Intermediate value

(샘플 표준화 인자)

(Sample normalization factor)

(표준화된 평균)

(Standardized average)

Model parameters

Transformed, standardized analysis value

Intermediate value

(샘플 표준화 인자)

(Sample normalization factor)

Transformed, standardized, replaced analysis values

은 감수성 지수 및 분류자 계산에 입력된다. Completed Nℓ xp sequence of values

Is entered into the sensitivity index and classifier calculations.

Susceptibility Index and Classifier

The following are the definitions of the analytical data and model parameters:

Justice

Analysis data

Justice

analysis Data

ℓ = reference set of samples (e.g., NHL cell line)

N _ℓ = sample size

p = number of probe pairs

x _ij = transformed, standardized analysis value for sample i, probe j

x _{ij '} = as described above using anti-correlation pair probe j' to probe j

Model parameters

β _ℓj = set ℓ coefficient for probe j

= 프로브 j에 대한 변환된 표준화된 분석 값의 세트 ℓ 평균

= Set ℓ average of transformed normalized analytical values for probe j

= 프로브 j에 대한 변환된 표준화된 분석 값의 세트 ℓ 평균

= Set ℓ average of transformed normalized analytical values for probe j

C _ℓ = classification split point

The equations for calculating the reference set model parameters, and the sensitivity index and classifier are presented below.

expression

Reference Set Model Parameters

Quotes and Classifiers

Clinical Trial 001 Results

Table 11 below provides sample accounting of the analyzed samples and clinical samples from clinical trial 001. 29 recording FFPE tumor samples from 24 DLBCL patients were submitted for qRT-PCR treatment. Three patients had multiple samples and all 24 patients had qRT-PCR results available for at least one sample. Of these 24, 21 had a sum of the product of tumor diameters (SPD) measurements reported at both baseline and at least one post-baseline visit.

Table 12 summarizes the pairwise spearman rank correlation between major genes and pair genes contributing to the sensitivity index. Based on cell line development samples, low expression genes in a particular group of patients would be expected to have, on average, relatively high expression of the corresponding pair, which is a ratio of self-standardized, and up-to-down-regulated expression pathways. (I.e. on a logarithmic scale of 2) provide an interpretation of the sensitivity index. The magnitude of the correlation between the pairs in the first clinical sample is statistically significant and significantly high, where the lower correlation estimate is -0.67 (P = 0.0004). These tests alone constitute an independent confirmation that the assay target sequences are expressed in tumor samples from the clinical population in vitro and the assay detects expression in the recorded FFPE tissue samples.

Table 13 summarizes the association between the measurements for each probe and the maximum decrease (or minimum increase) in post-baseline tumor SPD individually. Because the rank correlation is based on the difference (or ratio) between the post-baseline measurements versus the baseline measurements, a positive correlation means that higher expression of the probe is associated with tumor growth on average; A negative correlation means that higher expression of the probes is associated with tumor reduction on average. In particular, all major-pair probe pairs have a counter-directional association with the SPD. The P-value is consistent with the promising trend in the sample. All P-values are less than .5 (50% expected when there is no true association). All ranges are calculated as bootstrap 95th percentile confidence intervals based on 5,000 replicates sampled with replacements from DLBCL patient samples (N = 21). As sample size increases, narrower ranges will become available. Since no model-build or review is required to produce these results, they have a strong tendency to confirm that these qRT-PCR probe measurements are generally associated with a reduction in tumor SPD in patients treated with anti-CD40 Ab.1.

The multivariate sensitivity index is the weighted average of the probes in Tables 12 and 13. Since the weight in the cell line is not expected to reflect the optimal weight in the patient tumor sample, the weight in the cell line is limited to 1, -1 corresponding to the sign, equal-weighted mean, where the sign is assigned to IC25 in each probe and cell line. The association between resistance to anti-CD40 Ab.1 by For the clinical population, new weights are required. As a preliminary analysis based only on 21 samples, we decide to use the penalty multivariate regression procedure to select and estimate weights for the best 8 of 14 probes. These weights (coefficients) are shown in Table 14 and the association between the resulting susceptibility index and the SPD change from baseline is shown in FIG. 7. Larger multivariate sensitivity index values are associated with post-baseline SPD reduction (Spearman Rho = -0.58, P = 0.006). All ranges in Tables 13, 14 and 15 were calculated as Bootstrap 95th percentile confidence intervals based on 5,000 replicates sampled with replacements from DLBCL patient samples (N = 21). As sample size increases, narrower ranges will become available.

Using 26 samples from clinical trial 001, the ranges for μ _j and σ _j values obtained are shown in Table 15.

Clinical Trial 002 Results

Raw qRT-PCR results were successfully generated for 10 patients using recording samples. For these 10 patients, SPD changes from diagnosis, treatment group, multivariate sensitivity index, clinical response and baseline are shown in Table 16. Multivariate sensitivity index weights were taken from 21 clinical trial 001 patients (Table 14), thus these patients constitute a very small set of tests. Two of four patients with sensitivity index ≥ 0 showed some tumor contraction after anti-CD40 Ab.1 exposure, and four of six patients with sensitivity index <0 showed the best response of tumor increase or PD (SPD Although not available for the patient, the best clinical response results were available for that patient).

BCL6. qRT-PCR analysis includes a fifteenth probe for the BCL6 gene. Although not currently used in the multivariate sensitivity index, potential predictors of response to anti-CD40 Ab.1 have been previously identified. As shown in FIG. 8, although not significantly associated with SPD changes in the combined DLBCL patient sample (P = 0.25, N = 26), BCL6 tends to be lower in subjects with tumor growth (rho = -0.23) .

Although the foregoing invention has been described in more detail by way of illustration and example for purposes of clarity of understanding, the description and examples should not be construed as limiting the scope of the invention.

                                SEQUENCE LISTING <110> GENENTECH, INC.       DORNAN, David       BURINGTON, Bruce <120> METHODS AND COMPOSITIONS FOR ASSESSING       RESPONSIVENESS OF B-CELL LYMPHOMA TO TREATMENT WITH       ANTI-CD40 ANTIBODIES <130> 146392001740 <140> PCT / US2008 / 082920 <141> 2008-11-07 <150> US-60 / 986,277 <151> 2007-11-07 <160> 281 <170> FastSEQ for Windows Version 4.0 <210> 1 <211> 443 <212> PRT <213> Artificial Sequence <220> <223> Synthetic construct <400> 1 Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly  1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Tyr Ser Phe Thr Gly Tyr             20 25 30 Tyr Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val         35 40 45 Ala Arg Val Ile Pro Asn Ala Gly Gly Thr Ser Tyr Asn Gln Lys Phe     50 55 60 Lys Gly Arg Phe Thr Leu Ser Val Asp Asn Ser Lys Asn Thr Ala Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys                 85 90 95 Ala Arg Glu Gly Ile Tyr Trp Trp Gly Gln Gly Thr Leu Val Thr Val             100 105 110 Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser         115 120 125 Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys     130 135 140 Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu 145 150 155 160 Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu                 165 170 175 Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr             180 185 190 Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val         195 200 205 Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro     210 215 220 Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe 225 230 235 240 Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val                 245 250 255 Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe             260 265 270 Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro         275 280 285 Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr     290 295 300 Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val 305 310 315 320 Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala                 325 330 335 Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg             340 345 350 Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly         355 360 365 Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro     370 375 380 Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser 385 390 395 400 Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln                 405 410 415 Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His             420 425 430 Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly         435 440 <210> 2 <211> 219 <212> PRT <213> Artificial Sequence <220> <223> Synthetic construct <400> 2 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly  1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ser Ser Gln Ser Leu Val His Ser             20 25 30 Asn Gly Asn Thr Phe Leu His Trp Tyr Gln Gln Lys Pro Gly Lys Ala         35 40 45 Pro Lys Leu Leu Ile Tyr Thr Val Ser Asn Arg Phe Ser Gly Val Pro     50 55 60 Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile 65 70 75 80 Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr Tyr Phe Cys Ser Gln Thr                 85 90 95 Thr His Val Pro Trp Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys             100 105 110 Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu         115 120 125 Gln Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe     130 135 140 Tyr Pro Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln 145 150 155 160 Ser Gly Asn Ser Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser                 165 170 175 Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu             180 185 190 Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser         195 200 205 Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys     210 215 <210> 3 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 3 tgacaaaatg tagaggccat tca 23 <210> 4 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 4 catccgtctc ctctgcgata taa 23 <210> 5 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 5 ccgtcaaaca ccattt 16 <210> 6 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 6 ttgcaaggaa agaaattcaa acac 24 <210> 7 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 7 tgcttgaatc cattgactgc tt 22 <210> 8 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 8 acaacagcag taagaaga 18 <210> 9 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 9 caggtccctg cctttttaga ag 22 <210> 10 <211> 32 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 10 atcataaaga agagaagaga gacaagatta ag 32 <210> 11 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 11 agcctcatgg tctcat 16 <210> 12 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 12 ggccacagcc catcca 16 <210> 13 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 13 cttgccccta aatgttcctt tct 23 <210> 14 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 14 agtaactgac atgattagc 19 <210> 15 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 15 tttggaagtg aggcattgca 20 <210> 16 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 16 ccggagtccc cagagtcaa 19 <210> 17 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 17 agacgtacgt atcagcg 17 <210> 18 <211> 26 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 18 ctggaatgtg aagcgttata gaagat 26 <210> 19 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 19 ccttttttct ttcccaacac ttga 24 <210> 20 <211> 14 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 20 ctggcctcat ttct 14 <210> 21 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 21 cagcctctct tgtccctggt t 21 <210> 22 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 22 tccctagcaa tggacaaact ca 22 <210> 23 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 23 ccttatgtgt tgaatgtgg 19 <210> 24 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 24 gggatcctgt ttgccatcct 20 <210> 25 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 25 gcttcttggc cacctttttg 20 <210> 26 <211> 15 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 26 ttggtgctgg tcttt 15 <210> 27 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 27 ggcttcatag cattcgccta ct 22 <210> 28 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 28 tcacgtcgcc aaccatctt 19 <210> 29 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 29 cgtgaagtct agggacag 18 <210> 30 <211> 27 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 30 gacttgtatg tatgggagtg aggagtt 27 <210> 31 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 31 tctcttcaag ggcacagcta tg 22 <210> 32 <211> 14 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 32 cagggccatt gcaa 14 <210> 33 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 33 gccaaactgg aaacataaga gtga 24 <210> 34 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 34 gcatgacggt tcctgtgaaa 20 <210> 35 <211> 15 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 35 tgctcggtgg gatgg 15 <210> 36 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 36 cggaggttga ggtttttcct t 21 <210> 37 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 37 gacggttgaa tggcctctac a 21 <210> 38 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 38 tgtataagca cctactgaca aa 22 <210> 39 <211> 26 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 39 aggacttctt catgggtctt acagtt 26 <210> 40 <211> 27 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 40 aagtgacatt aaagacgatg tgtatgc 27 <210> 41 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 41 tgttagacca tgaaacatt 19 <210> 42 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 42 caggctgtgt tcttgcatct tg 22 <210> 43 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 43 gaccatgagg ctgcttctaa aaa 23 <210> 44 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 44 ctgcaaacag gtccct 16 <210> 45 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 45 ttggacccaa gggaaaactg 20 <210> 46 <211> 26 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 46 ggttaaaagt tgtggtttcc attctc 26 <210> 47 <211> 15 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 47 tggagacgca tttcg 15 <210> 48 <211> 25 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 48 gacatcccca ctcacgaata ttatg 25 <210> 49 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 49 ctgtcctttt ctgggctttc c 21 <210> 50 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 50 ccagtttctg cctctga 17 <210> 51 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 51 ggcatagagc agcactaaat gaca 24 <210> 52 <211> 26 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 52 ttctataacg cttcacattc cagatc 26 <210> 53 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 53 cactaaagaa acgatcagac 20 <210> 54 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 54 cgcactttgg ccttcctaga 20 <210> 55 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 55 tggaaggaga tgcagaagtc aga 23 <210> 56 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 56 cactgcttca taacctc 17 <210> 57 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 57 cctgcccagt cggcttct 18 <210> 58 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 58 gtccaagggt gacatttttc g 21 <210> 59 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 59 ctccaatgtg tcatctg 17 <210> 60 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 60 gggttactag tagccgccca ta 22 <210> 61 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 61 gcagggccag cattgc 16 <210> 62 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 62 caacctttgc actccac 17 <210> 63 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 63 tgtccatttt tttggctact ctga 24 <210> 64 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 64 cccaaacacc caggctctt 19 <210> 65 <211> 15 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 65 cagtgtggaa caatg 15 <210> 66 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 66 gctccagtgc cccaagatt 19 <210> 67 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 67 cgacggatcg cctctgaa 18 <210> 68 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 68 aaactgtgga tatcagcatg a 21 <210> 69 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 69 tgggcaactc agaaatactt cga 23 <210> 70 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 70 acgtcaatag gcacgtttgc t 21 <210> 71 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 71 ctcccaagat ataagaggc 19 <210> 72 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 72 gtccaccctc tcccctttct 20 <210> 73 <211> 26 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 73 cacgcactct agtacaaagc ataaga 26 <210> 74 <211> 15 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 74 ctcactccaa gaaac 15 <210> 75 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 75 cccaaaccga atcaccttaa ga 22 <210> 76 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 76 caggagggtg gccatcct 18 <210> 77 <211> 15 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 77 acagggctag ggcat 15 <210> 78 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 78 tctccatggc atcttcgtct t 21 <210> 79 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 79 atcccttacc ccaccctcaa 20 <210> 80 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 80 actcttaggc actttgg 17 <210> 81 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 81 cggcctcagg cacaagaa 18 <210> 82 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 82 gcagcccatc cagtgtcaat 20 <210> 83 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 83 atgtggacta tgtgatcct 19 <210> 84 <211> 27 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 84 catggtacat gagtggctat catactg 27 <210> 85 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 85 gtgagcacct tccttctttt tga 23 <210> 86 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 86 ctattatatg ggtttcagac aaa 23 <210> 87 <211> 27 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 87 gactattgtc tcctaaaccc aggacta 27 <210> 88 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 88 cccagtgcat ttaatgacca aa 22 <210> 89 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 89 agttccctcg tactgtc 17 <210> 90 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 90 atcaattttc ccgacgatct tc 22 <210> 91 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 91 cggttggcat ccatgtaaag t 21 <210> 92 <211> 14 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 92 tggctccaac actg 14 <210> 93 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 93 aggtccaccg tgatcaacat c 21 <210> 94 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 94 cagggaccag acgacatggt 20 <210> 95 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 95 acagcgagac ctccgt 16 <210> 96 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 96 caacttgtgg acggccagta 20 <210> 97 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 97 gtgccactga gggagaacat tt 22 <210> 98 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 98 aaactgcttc tacaagatt 19 <210> 99 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 99 cagcagagac cctgaaggaa a 21 <210> 100 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 100 caagccatga gttgccatca 20 <210> 101 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 101 aggtgcatat aagatctt 18 <210> 102 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 102 cccattctgc gtcatgctt 19 <210> 103 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 103 aatgcagttt agacacagcc aaac 24 <210> 104 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 104 tgttataact actccggaga cag 23 <210> 105 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 105 agttcagccc agatggaagg t 21 <210> 106 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 106 gcggcatcgc taaataagga 20 <210> 107 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 107 ttcagggaaa ggtgggc 17 <210> 108 <211> 29 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 108 cacagggact tgaagttgtt actaactaa 29 <210> 109 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 109 tgacgcagaa tgggatgaga 20 <210> 110 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 110 ctctctttgg gaatgtt 17 <210> 111 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 111 caaagcagcc agacgttgaa c 21 <210> 112 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 112 cacaccagat ccggaagaca 20 <210> 113 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 113 tttccctggg cgcagg 16 <210> 114 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 114 gggattccta ccccagattt cta 23 <210> 115 <211> 25 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 115 cagaaactgt tgttggactg catag 25 <210> 116 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 116 agtcagaaat gtaccaaaaa 20 <210> 117 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 117 tgagctgtag ctgcgtaagt acct 24 <210> 118 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 118 ggccttgtgc ctttcagaag 20 <210> 119 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 119 cttgatgcct gtcggc 16 <210> 120 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 120 tggctgccct acacatgct 19 <210> 121 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 121 caggatcccc tctaccactt tg 22 <210> 122 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 122 cctgctctat ctgcattt 18 <210> 123 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 123 gaggctcagc tgtgattgac at 22 <210> 124 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 124 cacccatatc ctcgaagcta gag 23 <210> 125 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 125 agaacatgga tgatacctc 19 <210> 126 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 126 tccagccaca gtcccctaga 20 <210> 127 <211> 27 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 127 tcctgaatgt tcctgatgat agtctct 27 <210> 128 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 128 agattaacat tgacagttcg aca 23 <210> 129 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 129 cgagaggaag gcgctgatc 19 <210> 130 <211> 26 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 130 acatcactcc atccttatac agcaaa 26 <210> 131 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 131 cctgcaagag attattt 17 <210> 132 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 132 ggatcctctt gacattcctc aaa 23 <210> 133 <211> 14 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <133> 133 ggccccccga tgga 14 <210> 134 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 134 ctccaccttt gaagacc 17 <210> 135 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 135 cgagggtgtg gccatatga 19 <210> 136 <211> 25 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 136 gaacaggcat tagaaatacc caaag 25 <210> 137 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 137 tgactagatg gctaatatg 19 <210> 138 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 138 ctactgcaag gcatgctttg at 22 <139> <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 139 tggccccctg cattga 16 <210> 140 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 140 tcccttcatc attgctg 17 <210> 141 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 141 dccggtgcag ttacacgtt 19 <210> 142 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 142 ccccaaaccc gtgacaac 18 <210> 143 <211> 15 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 143 cctccaagga gcctc 15 <210> 144 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 144 caaggccctc aacacattca 20 <210> 145 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 145 ggtacataac gggcatcttg atg 23 <210> 146 <211> 15 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 146 acctgttcgc ctttg 15 <210> 147 <211> 26 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 147 cctatgctgg agaaggatta gaaagt 26 <210> 148 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 148 cgatgattag aggtgcatgg aa 22 <210> 149 <211> 15 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 149 atgtggcaga taaag 15 <210> 150 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 150 tctcgccacc ctcaccat 18 <210> 151 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 151 gctgacagaa gtagatgcca ttgt 24 <210> 152 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 152 caaggcatcc ggtttg 16 <210> 153 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 153 aagtcgccct ggaacttcct 20 <210> 154 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 154 caccgagtcc tgctcctcat 20 <210> 155 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 155 atgagttgta cgagcagtc 19 <210> 156 <211> 25 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 156 catgagctgg tgaaaaatgg tattt 25 <210> 157 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 157 aaagctattc ctatcgtggc aaa 23 <210> 158 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 158 aaccagatac caagtttt 18 <210> 159 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 159 tccccagctc ttgccaaag 19 <210> 160 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 160 cagagaactc cctccaagtt gct 23 <210> 161 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 161 ctggagtaga aggacaacag 20 <210> 162 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 162 ggcaggccag ggtttgt 17 <210> 163 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 163 cgagatggct ggaaacacag a 21 <210> 164 <211> 14 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 164 aggcgctgtc tgtc 14 <210> 165 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 165 gactcagcct ctgggatgga 20 <210> 166 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 166 ggatccggaa gtagccattc t 21 <210> 167 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 167 tggattgtta aaaacagctg g 21 <210> 168 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 168 aggcggcttc ccatacct 18 <210> 169 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 169 cttcttccac cagcccaaaa 20 <210> 170 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 170 attgcaggaa agtacgcc 18 <210> 171 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 171 cccaaacctg caccactga 19 <210> 172 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 172 caagatgttg gcaaatgcaa a 21 <210> 173 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 173 ctgaaataca gcaaaaga 18 <210> 174 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 174 cctttgtggc atttattcat cagt 24 <210> 175 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 175 gcttctatga caagcagcct ttg 23 <210> 176 <211> 15 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 176 agggtgtccg attgg 15 <210> 177 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 177 ctctgtagca caggctggat tg 22 <210> 178 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 178 aggctgcagt gcaagattga 20 <210> 179 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 179 agtgcaatcc tgcaatt 17 <210> 180 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 180 ccacttggag gcctttcatc 20 <210> 181 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 181 aggttggcga tcaggaatac a 21 <210> 182 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 182 tcgggtgtgc tatgga 16 <210> 183 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 183 ccttgcctgg tttcgatgtt 20 <210> 184 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 184 gagcatttcc ctgtaggctt ctt 23 <210> 185 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 185 cccaaagcat aaaatt 16 <210> 186 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 186 caaccgttgg aaacataacc att 23 <210> 187 <211> 25 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 187 aacaatcagt agcacattgc atctg 25 <210> 188 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 188 agggagctgg gacact 16 <210> 189 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 189 tggactcact gaggctgacg ta 22 <210> 190 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 190 gattcccgag aacccttgat g 21 <210> 191 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 191 tcaccaagtt tgtgagttc 19 <210> 192 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 192 gctgccaatt ttgagcagtt t 21 <210> 193 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 193 gttcccagct tttccgttca 20 <210> 194 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 194 tgcaagaaag gatcaaa 17 <210> 195 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 195 tcttgcctgc cctgtgttg 19 <210> 196 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 196 tgccttcccc ttaataatgc a 21 <210> 197 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 197 aaaatgcggg tccctt 16 <210> 198 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 198 ctcccgctac acagaagtaa caaa 24 <210> 199 <211> 25 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 199 aaaacatccc tgctaccaat acatt 25 <210> 200 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 200 atggtagtca gttttgtatt tag 23 <210> 201 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 201 tccgttacaa gatgaggtct gtgt 24 <210> 202 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 202 dattctcctg gataacaacg ttga 24 <210> 203 <211> 13 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 203 tgctcacttc ccc 13 <210> 204 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 204 tccatccctt gacggttctg 20 <210> 205 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 205 agcccaagag gaatcaaaag atc 23 <206> 206 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 206 ccttcccaaa ctgcttt 17 <210> 207 <211> 27 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 207 gagtcatcac tgaggaagag aagaatt 27 <210> 208 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 208 tggcacgggc catacg 16 <210> 209 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 209 caaagccttc gctagtc 17 <210> 210 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 210 cctacacccc ttatccccat act 23 <210> 211 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 211 ccagggctat tggttgaatg a 21 <210> 212 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 212 ttattatcga aaccatcagc c 21 <210> 213 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 213 cgacctgcga gactcacaag 20 <210> 214 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 214 ggcacagcac tccgtctgt 19 <210> 215 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 215 aagctgacag agatacc 17 <210> 216 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 216 tctggctgtc ctttttataa tgca 24 <210> 217 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 217 cttggcaata gaacctggac aac 23 <210> 218 <211> 15 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 218 agtgagaact ttccc 15 <210> 219 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 219 gcaagaagaa gccactgaaa ca 22 <210> 220 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 220 gaaagcctta tcttcctcgt ccat 24 <210> 221 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 221 cccaagaagc aggcca 16 <210> 222 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 222 ggctgaaaat ggtggaaaag g 21 <210> 223 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 223 ctttgtccct gaggtgtcag ttt 23 <210> 224 <211> 15 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 224 ccaagatggc ggccg 15 <210> 225 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 225 tgtggatgag gcttccaaga a 21 <210> 226 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 226 cagcagggtc cggtcatact 20 <210> 227 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 227 agatcaaaga catcctcatc 20 <210> 228 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 228 ggcaggtgga ctacgagtca tac 23 <210> 229 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 229 gtctcctcgc tgccaggat 19 <210> 230 <211> 14 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 230 catggcggaa actg 14 <210> 231 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 231 ccggaacatt aagaccattg c 21 <210> 232 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 232 cccttggcag cattgatga 19 <210> 233 <211> 15 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 233 agtgcctggc agatg 15 <210> 234 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 234 ctgccacccc actcttaatc a 21 <210> 235 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 235 ggccaattga aacaaacagt tct 23 <210> 236 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 236 tggtggaaga acggtc 16 <210> 237 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 237 ggaagcctgc cacctcctat 20 <210> 238 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 238 tggcgcgagc attcttg 17 <210> 239 <211> 14 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 239 tgcggaccac catc 14 <210> 240 <211> 27 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 240 tgtccttgaa gcttgtatct gatatca 27 <210> 241 <211> 25 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 241 ttcaatacaa ggtcaaaatc agcaa 25 <210> 242 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 242 cactggattg tagaactt 18 <210> 243 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 243 agcctcagat gaaagaaaca atca 24 <210> 244 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 244 cacttgtgcc tgcagtttgg 20 <210> 245 <211> 15 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 245 aaccaggaaa aactc 15 <210> 246 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 246 aagcaggcga atcgtaatga g 21 <210> 247 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 247 tgcttgtgga atgtacagtg cat 23 <210> 248 <211> 13 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 248 cgtgcgccgc caa 13 <210> 249 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 249 cccttttctg ggtttgaagc t 21 <210> 250 <211> 26 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 250 ctgactgata caaagcacaa ttgaga 26 <210> 251 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 251 ctgtctctag aagtgcc 17 <210> 252 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 252 gctgtgaaag caacataaat ggat 24 <210> 253 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 253 ggcatgggaa cttaacagat gag 23 <210> 254 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 254 ttaaactgtc tacggttctt 20 <210> 255 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 255 cgctatccag aacctccact ct 22 <210> 256 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 256 caggtcatca cccttacttg ca 22 <210> 257 <211> 15 <212> DNA <213> Artificial Sequence <220> <223> Synthetic construct <400> 257 tcgacccctt tgctg 15 <210> 258 <211> 2034 <212> DNA <213> Homo sapiens <400> 258 aaaccttggc catggtcact tcctcttttc caatctctgt ggcagttttt gccctaataa 60 ccctgcaggt tggtactcag gacagtttta tagctgcagt gtatgaacat gctgtcattt 120 tgccaaataa aacagaaaca ccagtttctc aggaggatgc cttgaatctc atgaacgaga 180 atatagacat tctggagaca gcgatcaagc aggcagctga gcagggtgct cgaatcattg 240 tgactccaga agatgcactt tatggatgga aatttaccag ggaaactgtt ttcccttatc 300 tggaggatat cccagaccct caggtgaact ggattccgtg tcaagacccc cacagatttg 360 gtcacacacc agtacaagca agactcagct gcctggccaa ggacaactct atctatgtct 420 tggcaaattt gggggacaaa aagccatgta attcccgtga ctccacatgt cctcctaatg 480 gctactttca atacaatacc aatgtggtgt ataatacaga aggaaaactc gtggcacgtt 540 accataagta ccacctgtac tctgagcctc agtttaatgt ccctgaaaag ccggagttgg 600 tgactttcaa caccgcattt ggaaggtttg gcattttcac gtgctttgat atattcttct 660 atgatcctgg tgttaccctg gtgaaagatt tccatgtgga caccatactg tttcccacag 720 cttggatgaa cgttttgccc cttttgacag ctattgaatt ccattcagct tgggcaatgg 780 gaatgggagt taatcttctt gtggccaaca cacatcatgt cagcctaaat atgacaggaa 840 gtggtattta tgcaccaaat ggtcccaaag tgtatcatta tgacatgaag acagagttgg 900 gaaaacttct cctttcagag gtggattcac atcccctatc ctcgcttgcc tacccaacag 960 ctgttaattg gaatgcctac gccaccacca tcaaaccatt tccagtacag aaaaacactt 1020 tcaggggatt tatttccagg gatgggttca acttcacaga actttttgaa aatgcaggaa 1080 accttacagt ctgtcaaaag gagctttgct gtcatttaag ctacagaatg ttacaaaaag 1140 aagagaatga agtatacgtt ctaggagctt ttacaggatt acatggccga aggagaagag 1200 agtactggca ggtctgcaca atgctgaagt gcaaaactac taatttgaca acttgtggac 1260 ggccagtaga aactgcttct acaagatttg aaatgttctc cctcagtggc acatttggaa 1320 cagagtatgt ttttcctgaa gtgctactta ccgaaattca tctgtcacct ggaaaatttg 1380 aggtgctgaa agatgggcgt ttggtaaaca agaatggatc atctgggcct atactaacag 1440 tgtcactctt tgggaggtgg tacacaaagg actcacttta cagctcatgt gggaccagca 1500 attcagcaat aacttacctg ctaatattca tattattaat gatcatagct ttgcaaaata 1560 ttgtaatgtt atagggcgtc tctttatcac tcagcttctg catcatatgc ttggctgaat 1620 gtgtttatcg gcttcccaag tttactaaga aactttgaag ggctatttca gtagtataga 1680 ccagtgagtc ctaaatattt tttctcatca ataattattt tttaagtatt atgataatgt 1740 tgtccatttt tttggctact ctgaaatgtt gcagtgtgga acaatggaaa gagcctgggt 1800 gtttgggtca gataaatgaa gatcaaactc cagctccagc ctcatttgct tgagactttg 1860 tgtgtatggg ggacttgtat gtatgggagt gaggagtttc agggccattg caaacatagc 1920 tgtgcccttg aagagaatag taatgatggg aatttagagg tttatgactg aattcccttt 1980 gacattaaag actatttgaa ttcaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaa 2034 <210> 259 <211> 1498 <212> DNA <213> Homo sapiens <400> 259 gaggccagag tgccatcgaa ggtaattata gagacagtaa aatcctttta ctctgggaaa 60 aataaaatgc tgggtgtctc acaaaatttc agaacctgat ttcaaacgga tcataacaaa 120 gaggagatca aatttagcat ggtggactgc tcgacaggat atatttgtca atggaatgtt 180 tccacatatt ataccaccaa catgagaaaa aaatgatcat tgtttatttg aagcttgatg 240 atattctaac gctgcctttt ctcttctcat tttagagaaa aatgagcagg cggaattgtt 300 ggatttgtaa gatgtgcaga gatgaatcta agaggccccc ttcaaacctt actttggagg 360 aagtattaca gtgggcccag tcttttgaaa atttaatggc tacaaaatat ggtccagtag 420 tctatgcagc atatttaaaa atggagcaca gtgacgagaa tattcaattc tggatggcat 480 gtgaaaccta taagaaaatt gcctcacggt ggagcagaat ttctagggca aagaagcttt 540 ataagattta catccagcca cagtccccta gagagattaa cattgacagt tcgacaagag 600 agactatcat caggaacatt caggaaccca ctgaaacatg ttttgaagaa gctcagaaaa 660 tagtctatat gcatatggaa agggattcct accccagatt tctaaagtca gaaatgtacc 720 aaaaactttt gaaaactatg cagtccaaca acagtttctg actacaactc aaaagtttaa 780 atagaaaaca gtatattgaa agtggtgggt ttgatctttt tatttagaaa cccacaaaat 840 cagaaacaca gtacaaataa aacagaaatc aaactataag ttgactttta gttcctaaaa 900 agaaacatat ttcaaaagca atggaatcta gaattcttat aacatgaata acaaaatgta 960 cagcaagcct atgtagttca attaatatat aaggaaaagg aaggtctttc ttcatgatac 1020 aagcattata aagtttttac tgtagtagtc aattaatgga tatttccttg ttaataaaat 1080 tttgtgtcat aatttacaaa ttagttcttt aaaaattgtt gttatatgaa ttgtgtttct 1140 agcatgaatg ttctatagag tactctaaat aacttgaatt tatagacaaa tgctactcac 1200 agtacaatca attgtattat accatgagaa aatcaaaaag gtgttcttca gagacatttt 1260 atctataaaa ttttcctact attatgttca ttaacaaact tctttatcac atgtatcttc 1320 tacatgtaaa acatttctga tgatttttta acaaaaaata tatgaatttc ttcatttgct 1380 cttgcatcta cattgctata aggatataaa atgtggtttc tatattttga gatgtttttt 1440 ccttacaatg tgaactcatc gtgatcttgg aaatcaataa agtcaaatat caactaaa 1498 <210> 260 <211> 3260 <212> DNA <213> Homo sapiens <400> 260 ccatcccata gtgagggaag acacgcggaa acaggcttgc acccagacac gacaccatgc 60 atctcctcgg cccctggctc ctgctcctgg ttctagaata cttggctttc tctgactcaa 120 gtaaatgggt ttttgagcac cctgaaaccc tctacgcctg ggagggggcc tgcgtctgga 180 tcccctgcac ctacagagcc ctagatggtg acctggaaag cttcatcctg ttccacaatc 240 ctgagtataa caagaacacc tcgaagtttg atgggacaag actctatgaa agcacaaagg 300 atgggaaggt tccttctgag cagaaaaggg tgcaattcct gggagacaag aataagaact 360 gcacactgag tatccacccg gtgcacctca atgacagtgg tcagctgggg ctgaggatgg 420 agtccaagac tgagaaatgg atggaacgaa tacacctcaa tgtctctgaa aggccttttc 480 cacctcatat ccagctccct ccagaaattc aagagtccca ggaagtcact ctgacctgct 540 tgctgaattt ctcctgctat gggtatccga tccaattgca gtggctccta gagggggttc 600 caatgaggca ggctgctgtc acctcgacct ccttgaccat caagtctgtc ttcacccgga 660 gcgagctcaa gttctcccca cagtggagtc accatgggaa gattgtgacc tgccagcttc 720 aggatgcaga tgggaagttc ctctccaatg acacggtgca gctgaacgtg aagcacaccc 780 cgaagttgga gatcaaggtc actcccagtg atgccatagt gagggagggg gactctgtga 840 ccatgacctg cgaggtcagc agcagcaacc cggagtacac gacggtatcc tggctcaagg 900 atgggacctc gctgaagaag cagaatacat tcacgctaaa cctgcgcgaa gtgaccaagg 960 accagagtgg gaagtactgc tgtcaggtct ccaatgacgt gggcccggga aggtcggaag 1020 aagtgttcct gcaagtgcag tatgccccgg aaccttccac ggttcagatc ctccactcac 1080 cggctgtgga gggaagtcaa gtcgagtttc tttgcatgtc actggccaat cctcttccaa 1140 caaattacac gtggtaccac aatgggaaag aaatgcaggg aaggacagag gagaaagtcc 1200 acatcccaaa gatcctcccc tggcacgctg ggacttattc ctgtgtggca gaaaacattc 1260 ttggtactgg acagaggggc ccgggagctg agctggatgt ccagtatcct cccaagaagg 1320 tgaccacagt gattcaaaac cccatgccga ttcgagaagg agacacagtg accctttcct 1380 gtaactacaa ttccagtaac cccagtgtta cccggtatga atggaaaccc catggcgcct 1440 gggaggagcc atcgcttggg gtgctgaaga tccaaaacgt tggctgggac aacacaacca 1500 tcgcctgcgc acgttgtaat agttggtgct cgtgggcctc ccctgtcgcc ctgaatgtcc 1560 agtatgcccc ccgagacgtg agggtccgga aaatcaagcc cctttccgag attcactctg 1620 gaaactcggt cagcctccaa tgtgacttct caagcagcca ccccaaagaa gtccagttct 1680 tctgggagaa aaatggcagg cttctgggga aagaaagcca gctgaatttt gactccatct 1740 ccccagaaga tgctgggagt tacagctgct gggtgaacaa ctccatagga cagacagcgt 1800 ccaaggcctg gacacttgaa gtgctgtatg cacccaggag gctgcgtgtg tccatgagcc 1860 cgggggacca agtgatggag gggaagagtg caaccctgac ctgtgagagt gacgccaacc 1920 ctcccgtctc ccactacacc tggtttgact ggaataacca aagcctcccc caccacagcc 1980 agaagctgag attggagccg gtgaaggtcc agcactcggg tgcctactgg tgccagggga 2040 ccaacagtgt gggcaagggc cgttcgcctc tcagcaccct tactgtctac tatagcccgg 2100 agaccatcgg caggcgagtg gctgtgggac tcgggtcctg cctcgccatc ctcatcctgg 2160 caatctgtgg gctcaagctc cagcgacgtt ggaagaggac acagagccag caggggcttc 2220 aggagaattc cagcggccag agcttctttg tgaggaataa aaaggttaga agggcccccc 2280 tctctgaagg cccccactcc ctgggatgct acaatccaat gatggaagat ggcattagct 2340 acaccaccct gcgctttccc gagatgaaca taccacgaac tggagatgca gagtcctcag 2400 agatgcagag acctccccgg acctgcgatg acacggtcac ttattcagca ttgcacaagc 2460 gccaagtggg cgactatgag aacgtcattc cagattttcc agaagatgag gggattcatt 2520 actcagagct gatccagttt ggggtcgggg agcggcctca ggcacaagaa aatgtggact 2580 atgtgatcct caaacattga cactggatgg gctgcagcag aggcactggg ggcagcgggg 2640 gccagggaag tccccgagtt tccccagaca ccgccacatg gcttcctcct gcgtgcatgt 2700 gcgcacacac acacacacac gcacacacac acacacacac tcactgcgga gaaccttgtg 2760 cctggctcag agccagtctt tttggtgagg gtaaccccaa acctccaaaa ctcctgcccc 2820 tgttctcttc cactctcctt gctacccaga aatcatctaa atacctgccc tgacatgcac 2880 acctcccctg ccccaccagc ccactggcca tctccacccg gagctgctgt gtcctctgga 2940 tctgctcgtc attttccttc ccttctccat ctctctggcc ctctacccct gatctgacat 3000 ccccactcac gaatattatg cccagtttct gcctctgagg gaaagcccag aaaaggacag 3060 aaacgaagta gaaaggggcc cagtcctggc ctggcttctc ctttggaagt gaggcattgc 3120 acggggagac gtacgtatca gcggcccctt gactctgggg actccgggtt tgagatggac 3180 acactggtgt ggattaacct gccagggaga cagagctcac aataaaaatg gctcagatgc 3240 cacttcaaag aaaaaaaaaa 3260 <210> 261 <211> 2433 <212> DNA <213> Homo sapiens <400> 261 atgattccgg tgacagagct ccgctacttt gcggacacgc agccagcata ccggatcctg 60 aagccgtggt gggatgtgtt cacagactac atctctatcg tcatgctgat gattgccgtc 120 ttcgggggga cgctgcaggt cacccaagac aagatgatct gcctgccttg taagtgggtc 180 accaaggact cctgcaatga ttcgttccgg ggctgggcag cccctggccc ggagcccacc 240 taccccaact ccaccattct gccgacccct gacacgggcc ccacaggcat caagtatgac 300 ctggaccggc accagtacaa ctacgtggac gctgtgtgct atgagaaccg actgcactgg 360 tttgccaagt acttccccta cctggtgctt ctgcacacgc tcatcttcct ggcctgcagc 420 aacttctggt tcaaattccc gcgcaccagc tcgaagctgg agcactttgt gtctatcctg 480 ctgaagtgct tcgactcgcc ctggaccacg agggccctgt cggagacagt ggtggaggag 540 agcgacccca agccggcctt cagcaagatg aatgggtcca tggacaaaaa gtcatcgacc 600 gtcagtgagg acgtggaggc caccgtgccc atgctgcagc ggaccaagtc acggatcgag 660 cagggtatcg tggaccgctc agagacgggc gtgctggaca agaaggaggg ggagcaagcc 720 aaggcgctgt ttgagaaggt gaagaagttc cggacccatg tggaggaggg ggacattgtg 780 taccgcctct acatgcggca gaccatcatc aaggtgatca agttcatcct catcatctgc 840 tacaccgtct actacgtgca caacatcaag ttcgacgtgg actgcaccgt ggacattgag 900 agcctgacgg gctaccgcac ctaccgctgt gcccaccccc tggccacact cttcaagatc 960 ctggcgtcct tctacatcag cctagtcatc ttctacggcc tcatctgcat gtatacactg 1020 tggtggatgc tacggcgctc cctcaagaag tactcgtttg agtcgatccg tgaggagagc 1080 agctacagcg acatccccga cgtcaagaac gacttcgcct tcatgctgca cctcattgac 1140 caatacgacc cgctctactc caagcgcttc gccgtcttcc tgtcggaggt gagtgagaac 1200 aagctgcggc agctgaacct caacaacgag tggacgctgg acaagctccg gcagcggctc 1260 accaagaacg cgcaggacaa gctggagctg cacctgttca tgctcagtgg catccctgac 1320 actgtgtttg acctggtgga gctggaggtc ctcaagctgg agctgatccc cgacgtgacc 1380 atcccgccca gcattgccca gctcacgggc ctcaaggagc tgtggctcta ccacacagcg 1440 gccaagattg aagcgcccgc gctggccttc ctgcgcgaga acctgcgggc gctgcacatc 1500 aagttcaccg acatcaagga gatcccgctg tggatctata gcctgaagac actggaggag 1560 ctgcacctga cgggcaacct gagcgcggag aacaaccgct acatcgtcat cgacgggctg 1620 cgggagctca aacgcctcaa ggtgctgcgg ctcaagagca acctaagcaa gctgccacag 1680 gtggtcacag atgtgggcgt gcacctgcag aagctgtcca tcaacaatga gggcaccaag 1740 ctcatcgtcc tcaacagcct caagaagatg gcgaacctga ctgagctgga gctgatccgc 1800 tgtgacctgg agcgcatccc ccactccatc ttcagcctcc acaacctgca ggagattgac 1860 ctcaaggaca acaacctcaa gaccatcgag gagatcatca gcttccagca cctgcaccgc 1920 ctcacctgcc ttaagctgtg gtacaaccac atcgcctaca tccccatcca gatcggcaac 1980 ctcaccaacc tggagcgcct ctacctgaac cgcaacaaga tcgagaagat ccccacccag 2040 ctcttctact gccgcaagct gcgctacctg gacctcagcc acaacaacct gaccttcctc 2100 cctgccgaca tcggcctcct gcagaacctc cagaacctag ccatcacggc caaccggatc 2160 gagacgctcc ctccggagct cttccagtgc cggaagctgc gggccctgca cctgggcaac 2220 aacgtgctgc agtcactgcc ctccagggtg ggcgagctga ccaacctgac gcagatcgag 2280 ctgcggggca accggctgga gtgcctgcct gtggagctgg gcgagtgccc actgctcaag 2340 cgcagcggct tggtggtgga ggaggacctg ttcaacacac tgccacccga ggtgaaggag 2400 cggctgtgga gggctgacaa ggagcaggcc tga 2433 <210> 262 <211> 1616 <212> DNA <213> Homo sapiens <400> 262 gccaaggctg gggcagggga gtcagcagag gcctcgctcg ggcgcccagt ggtcctgccg 60 cctggtctca cctcgctatg gttcgtctgc ctctgcagtg cgtcctctgg ggctgcttgc 120 tgaccgctgt ccatccagaa ccacccactg catgcagaga aaaacagtac ctaataaaca 180 gtcagtgctg ttctttgtgc cagccaggac agaaactggt gagtgactgc acagagttca 240 ctgaaacgga atgccttcct tgcggtgaaa gcgaattcct agacacctgg aacagagaga 300 cacactgcca ccagcacaaa tactgcgacc ccaacctagg gcttcgggtc cagcagaagg 360 gcacctcaga aacagacacc atctgcacct gtgaagaagg ctggcactgt acgagtgagg 420 cctgtgagag ctgtgtcctg caccgctcat gctcgcccgg ctttggggtc aagcagattg 480 ctacaggggt ttctgatacc atctgcgagc cctgcccagt cggcttcttc tccaatgtgt 540 catctgcttt cgaaaaatgt cacccttgga caagctgtga gaccaaagac ctggttgtgc 600 aacaggcagg cacaaacaag actgatgttg tctgtggtcc ccaggatcgg ctgagagccc 660 tggtggtgat ccccatcatc ttcgggatcc tgtttgccat cctcttggtg ctggtcttta 720 tcaaaaaggt ggccaagaag ccaaccaata aggcccccca ccccaagcag gaaccccagg 780 agatcaattt tcccgacgat cttcctggct ccaacactgc tgctccagtg caggagactt 840 tacatggatg ccaaccggtc acccaggagg atggcaaaga gagtcgcatc tcagtgcagg 900 agagacagtg aggctgcacc cacccaggag tgtggccacg tgggcaaaca ggcagttggc 960 cagagagcct ggtgctgctg ctgctgtggc gtgagggtga ggggctggca ctgactgggc 1020 atagctcccc gcttctgcct gcacccctgc agtttgagac aggagacctg gcactggatg 1080 cagaaacagt tcaccttgaa gaacctctca cttcaccctg gagcccatcc agtctcccaa 1140 cttgtattaa agacagaggc agaagtttgg tggtggtggt gttggggtat ggtttagtaa 1200 tatccaccag accttccgat ccagcagttt ggtgcccaga gaggcatcat ggtggcttcc 1260 ctgcgcccag gaagccatat acacagatgc ccattgcagc attgtttgtg atagtgaaca 1320 actggaagct gcttaactgt ccatcagcag gagactggct aaataaaatt agaatatatt 1380 tatacaacag aatctcaaaa acactgttga gtaaggaaaa aaaggcatgc tgctgaatga 1440 tgggtatgga actttttaaa aaagtacatg cttttatgta tgtatattgc ctatggatat 1500 atgtataaat acaatatgca tcatatattg atataacaag ggttctggaa gggtacacag 1560 aaaacccaca gctcgaagag tggtgacgtc tggggtgggg aagaagggtc tggggg 1616 <210> 263 <211> 733 <212> DNA <213> Homo sapiens <400> 263 aaacagcagg aaatagaaac ttaagagaaa tacacacttc tgagaaactg aaacgacagg 60 ggaaaggagg tctcactgag caccgtccca gcatccggac accacagcgg cccttcgctc 120 cacgcagaaa accacacttc tcaaaccttc actcaacact tccttcccca aagccagaag 180 atgcacaagg aggaacatga ggtggctgtg ctgggggcac cccccagcac catccttcca 240 aggtccaccg tgatcaacat ccacagcgag acctccgtgc ccgaccatgt cgtctggtcc 300 ctgttcaaca ccctcttctt gaactggtgc tgtctgggct tcatagcatt cgcctactcc 360 gtgaagtcta gggacaggaa gatggttggc gacgtgaccg gggcccaggc ctatgcctcc 420 accgccaagt gcctgaacat ctgggccctg attctgggca tcctcatgac cattggattc 480 atcctgttac tggtattcgg ctctgtgaca gtctaccata ttatgttaca gataatacag 540 gaaaaacggg gttactagta gccgcccata gcctgcaacc tttgcactcc actgtgcaat 600 gctggccctg cacgctgggg ctgttgcccc tgcccccttg gtcctgcccc tagatacagc 660 agtttatacc cacacacctg tctacagtgt cattcaataa agtgcacgtg cttgtgaaaa 720 aaaaaaaaaa aaa 733 <210> 264 <211> 8787 <212> DNA <213> Homo sapiens <400> 264 ggccgcagct ccccggcgga ggcaagaggt ggttgggggg gaccatggct gacgttttcc 60 cgggcaacga ctccacggcg tctcaggacg tggccaaccg cttcgcccgc aaaggggcgc 120 tgaggcagaa gaacgtgcac gaggtgaagg accacaaatt catcgcgcgc ttcttcaagc 180 agcccacctt ctgcagccac tgcaccgact tcatctgggg gtttgggaaa caaggcttcc 240 agtgccaagt ttgctgtttt gtggtccaca agaggtgcca tgaatttgtt actttttctt 300 gtccgggtgc ggataaggga cccgacactg atgaccccag gagcaagcac aagttcaaaa 360 tccacactta cggaagcccc accttctgcg atcactgtgg gtcactgctc tatggactta 420 tccatcaagg gatgaaatgt gacacctgcg atatgaacgt tcacaagcaa tgcgtcatca 480 atgtccccag cctctgcgga atggatcaca ctgagaagag ggggcggatt tacctaaagg 540 ctgaggttgc tgatgaaaag ctccatgtca cagtacgaga tgcaaaaaat ctaatcccta 600 tggatccaaa cgggctttca gatccttatg tgaagctgaa acttattcct gatcccaaga 660 atgaaagcaa gcaaaaaacc aaaaccatcc gctccacact aaatccgcag tggaatgagt 720 cctttacatt caaattgaaa ccttcagaca aagaccgacg actgtctgta gaaatctggg 780 actgggatcg aacaacaagg aatgacttca tgggatccct ttcctttgga gtttcggagc 840 tgatgaagat gccggccagt ggatggtaca agttgcttaa ccaagaagaa ggtgagtact 900 acaacgtacc cattccggaa ggggacgagg aaggaaacat ggaactcagg cagaaattcg 960 agaaagccaa acttggccct gctggcaaca aagtcatcag tccctctgaa gacaggaaac 1020 aaccttccaa caaccttgac cgagtgaaac tcacggactt caatttcctc atggtgttgg 1080 gaaaggggag ttttggaaag gtgatgcttg ccgacaggaa gggcacagaa gaactgtatg 1140 caatcaaaat cctgaagaag gatgtggtga ttcaggatga tgacgtggag tgcaccatgg 1200 tagaaaagcg agtcttggcc ctgcttgaca aacccccgtt cttgacgcag ctgcactcct 1260 gcttccagac agtggatcgg ctgtacttcg tcatggaata tgtcaacggt ggggacctca 1320 tgtaccacat tcagcaagta ggaaaattta aggaaccaca agcagtattc tatgcggcag 1380 agatttccat cggattgttc tttcttcata aaagaggaat catttatagg gatctgaagt 1440 tagataacgt catgttggat tcagaaggac atatcaaaat tgctgacttt gggatgtgca 1500 aggaacacat gatggatgga gtcacgacca ggaccttctg tgggactcca gattatatcg 1560 ccccagagat aatcgcttat cagccgtatg gaaaatctgt ggactggtgg gcctatggcg 1620 tcctgttgta tgaaatgctt gccgggcagc ctccatttga tggtgaagat gaagacgagc 1680 tatttcagtc tatcatggag cacaacgttt cctatccaaa atccttgtcc aaggaggctg 1740 tttctatctg caaaggactg atgaccaaac acccagccaa gcggctgggc tgtgggcctg 1800 agggggagag ggacgtgaga gagcatgcct tcttccggag gatcgactgg gaaaaactgg 1860 agaacaggga gatccagcca ccattcaagc ccaaagtgtg tggcaaagga gcagagaact 1920 ttgacaagtt cttcacacga ggacagcccg tcttaacacc acctgatcag ctggttattg 1980 ctaacataga ccagtctgat tttgaagggt tctcgtatgt caacccccag tttgtgcacc 2040 ccatcttaca gagtgcagta tgaaactcac cagcgagaac aaacacctcc ccagccccca 2100 gccctccccg cagtgggaag tgaatcctta accctaaaat tttaaggcca cggccttgtg 2160 tctgattcca tatggaggcc tgaaaattgt agggttatta gtccaaatgt gatcaactgt 2220 tcagggtctc tctcttacaa ccaagaacat tatcttagtg gaagatggta cgtcatgctc 2280 agtgtccagt ttaattctgt agaagttacg tctggctcta ggttaaccct tcctagaaag 2340 caagcagact gttgccccat tttgggtaca atttgatata ctttccatac cctccatctg 2400 tggatttttc agcattggaa tcccccaacc agagatgtta aagtgagcct gtcccaggaa 2460 acatctccac ccaagacgtc tttggaatcc aagaacagga agccaagaga gtgagcaggg 2520 agggattggg ggtgggggag gcctcaaaat accgactgcg tccattctct gcctccatgg 2580 aaacagcccc tagaatctga aaggccggga taaacctaat cactgttccc aaacattgac 2640 aaatcctaac ccaaccatgg tccagcagtt accagtttaa acaaaaaaac ctcagatgag 2700 tgttgggtga atctgtcatc tggtaccctc cttggttgat aactgtcttg atacttttca 2760 ttctttgtaa gaggccaaat cgtctaagga cgttgctgaa caagcgtgtg aaatcatttc 2820 agatcaagga taagccagtg tgtacatatg ttcattttaa tctctgggag attatttttc 2880 catccagggt gccatcagta atcatgccac tactcaccag tgttgttcgc caacacccac 2940 ccccacacac accaacattt tgctgcctac cttgttatcc ttctcaagaa gctgaagtgt 3000 acgccctctc cccttttgtg cttatttatt taataggctg cagtgtcgct tatgaaagta 3060 cgatgtacag taacttaatg gaagtgctga ctctagcatc agcctctacc gattgatttt 3120 cctcccttct ctagccctgg atgtccactt agggataaaa agaatatggt tttggttccc 3180 atttctagtt cacgttgaat gacaggcctg gagctgtaga atcaggaaac ccggatgcct 3240 aacagctcaa agatgttttg ttaatagaag gattttaata cgttttgcaa atgcatcatg 3300 caatgaattt tgcatgttta taataaacct taataacaag tgaatctata ttattgatat 3360 aatcgtatca agtataaaga gagtattata ataattttat aagacacaat tgtgctctat 3420 ttgtgcaggt tcttgtttct aatcctcttt tctaattaag ttttagctga atcccttgct 3480 tctgtgcttt ccctccctgc acatgggcac tgtatcagat agattacttt ttaaatgtag 3540 ataaaatttc aaaaatgaat ggctagttta cgtgatagat taggctctta ctacatatgt 3600 gtgtgtatat atatgtattt gattctacct gcaaacaaat ttttattggt gaggactatt 3660 tttgagctga cactccctct tagtttcttc atgtcacctt tcgtcctggt tcctccgcca 3720 ctcttcctct tggggacaac aggaagtgtc tgattccagt ctgcctagta cgttggtaca 3780 cacgtggcat tgccgcagca cctgggctga cctttgtgtg tgcgtgtgtg tgtgtttcct 3840 tcttcccttc agcctgtgac tgttgctgac tccaggggtg ggagggatgg ggagactccc 3900 ctcttgctgt gtgtactgga cacgcaggaa gcatgctgtc ttgctgcctc tgcaacgacc 3960 tgtcgtttgc tccagcatgc acaaacttcg tgagaccaac acagccgtgc cctgcaggca 4020 ccagcacgtg cttttcagag gctgcggact ttcttccagc cattgtggca ttggcctttc 4080 cagtcttggg aggagcgcgc tgctttggtg agacaccccc atgcaaggtc ctcagagtag 4140 ccgggttcta ccacaaacag aaacagaatg aaagtagctg tcagtccttg tagagagccg 4200 ctctgtttcc tcccagaagc atctcccagc taagctcgca ttatttttct cctctggctg 4260 tttgcctgaa gttcacagaa cacacaacca tgaaaggctt tttgaggtga gaggcccagg 4320 tggtcctggc aaccctgagt agaaggagag acggggtagg gaacgggccc ggccagaaaa 4380 gaaccatttc ttctgccatc ttttatgcac catagacatc gagactccag ggggtcctgg 4440 ctcccctgtc cctgcagccc tgcaggtcag tgcatgatct gggttcgtgt cctgaccagg 4500 tgctcctcct ttgatccgag gggaaaggga ctggtttata gaaagagcct aggagacaaa 4560 agggccagtc cccctgccca gaatggagca gcagcaggac agacccccac gaggcccccc 4620 agagaggagg aagatcccac ggaggaacac atgaggttag ggacccttgt tcagcacccc 4680 aaacagcctg cctgtttaaa gcaggcagca ggcttaggcc ttccctgcaa ccccaacacc 4740 cacaagtttg tttctctagg aaacacattc actgtctcag ctggctgtta ctctctcaga 4800 ccatatggca aagttttcca agaaaatgcc ccgacagggg tgcccagcac actgcctgag 4860 ggacaacaga catcagaaca aacccccaga gagaaacagt caaaatcagg gcccggtgca 4920 gtgttgtcat gtggaacctg ctttatccat tgctgagtgt tgaatgtggg taatggttag 4980 ggctttccag atctcagcag ccaaagacag ttattgttgg aagactgtca tgtagataac 5040 catgagcaat ggctcgcctc agaatcagtt cataaaattc tatggtactg gccccttcgt 5100 gggtattgtg tgaaatgaga tggtggcgag gggtgcgctg tggaactgcc gcagccacgc 5160 aggaggtccc tgggggatgc tttgggaagt ccttgcccct gagcactgcc tgattgccag 5220 ggcctgtgga ggtctaggcc gcctggcaga atctagcacc gtccgaatcc ccgcaggacc 5280 catggagcta tgaccacacc aggccattca aatggctctg cattatcttc ccttggaagg 5340 tggccactcc tcggtggcag ggcctttccc tgaggctgca ggccgtgggc tggcagcccg 5400 tctcttggca tttcaattga aggtcaccag gtgctgggtt tgaaaggaag tcactggagt 5460 gctgccaggg gccgccctcc aaggttaatg agaggcccac atccaggcaa gaactaattc 5520 aaaaggcaga tcagaaacca caggagtcaa aattattgct ccggcagtgc ttcccttcct 5580 ttcatccact ggcctcgtgt ggtccatgca gggccactgt ctgccctttc tgatgccacg 5640 tattaggctt tcttactcag aattttgata gaaaaccatg gggccaagag ctctggaagc 5700 ctggccggaa agaccaaggt tcatgcagcc caacaaatga ttgttgagca cctctcggag 5760 ccaaagtcct taggcgagtg tggtgacttc ctggaaggag gatgcagact tccagagagc 5820 ccccccaacg gacgtgctga gaagggagag ggaggcgggg gctgtagtca ggaaggagcc 5880 agagaagaac agggtttggg tgcatccaga aatatgcctg cagtaggagg gagaggaagg 5940 ggtgccaccg tcaacggctt cccatcggag gtggttggtg cagatggaag tttctgtctg 6000 ctggccctca agagagtgtt ttgccaggga cacagtctgt tcctcctcag aaaacacccc 6060 ccaaatgcta acaacatccc caccagctgc tagaagcccc tttcccctcc ccaccttgaa 6120 gtagctcata gttctctggg cagagccaga ccatccagtg taccccagag gccagtaggt 6180 tcctgcccat tttcctctct ggcttcctgc caagaattat ggcagctgag gatgaatgga 6240 gaagtaaaaa caactaacac cgcacaacta acaactaaca ccgcagttcc cacctgggtt 6300 ccacttagca ggagacattt cggagggttt tttttgtttt tgttcctgtt tttttttttt 6360 ttgctggaat ttgttttctc agtactgaaa agagaaaaag tgacaatctt gtatttttaa 6420 aagcctcgga aaggtgatac catctgacag tcattttctc acgttggtct tctaaagtca 6480 cctatttctt gtgtgtgcac atcacaccat ttcctgtttc tttataaccc gacaagggta 6540 ggagtgcctg tttcccctgc tgggcacacc agacaatcgt aatcacaaaa cagacactga 6600 gccaggggcc caaagggtgt gatcatgaga gttaccggga cagcagtagg catgacagtc 6660 accaggaagg acaagggtgc tctgttgtta gtggccacac accaatttga caaggagtgt 6720 tgcgaaattt ttatttattt atttatttat tttgagatgg agtttcactc ttgttgccca 6780 ggctggagtg cggtggtaca atctcggctc actgcaacct ccacctccca ggttcaagcg 6840 attctcctgc ctcagcctcc caagtacctg ggactacagg tgcgtgccac cacacccagc 6900 taaattttgt gtttttagta gagatggggt ttcaccatgt tggccaggat ggtcttgaac 6960 ccctgacctc atgatctgcc tgcctcggcc tcccaaagtg ctgggattac aggcatgagc 7020 caccacgccc agccaaaata tttttttaaa gtcattttcc ttaagctgct tgggctacat 7080 gtgaaataca ctggacggtc aacattcctg tctcctccca tttgggctga tgcagcagat 7140 ccagggaatg ttacctgttt ctgctgctag aagatccagg aaattgggaa ggttacctga 7200 cgcacacatg gatgaaggcc atcatctaga aatggggtca accacaattg tgttaattcc 7260 gtagtgtcag ggattcttcg ggaaggtcaa cagtatgaag gattctgacc cctgtgcctc 7320 ccatttatgt gatcaggtga cagttaataa ccgtggaggt cacactcagc catccaacag 7380 ccttacagtg accctacaca aaagccccca aattccaaag actttttctt aacctaaagg 7440 aagaaattat ttgttaattc cagtagagca actgaatata ctgggctatt tgtacttttt 7500 tatagagaac tttaataata attctttaaa aatgagtttt tagaacaaag caactgacga 7560 tttcctaaga ttccaatgcc ctggagcttg taggaggact tagcctgggt cagctggagc 7620 acccccgacc tgatctccca ctgccagatt ttcccatgct cctagggtat ggagtccacg 7680 tgggaatgac tgcaagttca ggtggaactt ggccgactga tgctctgcga gtttttaata 7740 gacactgggg acaactgctt aaggtttaga aacttccaaa ccacaggaaa gacattttta 7800 gtgtccccca tccagaggca gccctggaat aggattccca ggggtttctg ggaccccttt 7860 ccttgctccg tgaggctctg tggccatctt ttggcaggag gaggatgctt ccttggctct 7920 gtgcccagac ccgcctggtc cccaggtctc tcaccttggg tgaagattca gagatgccct 7980 gtaaggattt tgcccactgg gcaactcaga aatacttcga tctcccaaga tataagaggc 8040 agcagcaaac gtgcctattg acgtctgttt catagttacc acttacgcga gtagacagaa 8100 ctcggctttt cagaaaatag gtgtcaagtc cactttataa gaaccttttt ttctaaaata 8160 agataaaagg tggctttgca ttttctgatt aaacgactgt gtctttgtca cctctgctta 8220 actttaggag tatccattcc tgtgattgta gacttttgtt gatattcttc ctggaagaat 8280 atcattcttt tcttgaaggg ttggtttact agaatattca aaatcaatca tgaaggcagt 8340 tactattttg agtctaaagg ttttctaaaa attaacctca catcccttct gttagggtct 8400 ttcagaatat cttttataaa cagaagcatt tgaagtcatt gcttttgcta catgatttgt 8460 gtgtgtgaag gacataccac gtttaaatca ttaattgaaa aacatcatat aagccccaac 8520 tttgtttgga ggaagagacg gaggttgagg tttttccttc tgtataagca cctactgaca 8580 aaatgtagag gccattcaac cgtcaaacac catttggtta tatcgcagag gagacggatg 8640 tgtaaattac tgcattgctt tttttttcag tttgtataac ctctaatctc cgtttgcatg 8700 atacgctttg ttagaaacat taattgtagt ttggaagcaa gtgtgtatga ataaagataa 8760 tgatcattcc aaaaaaaaaa aaaaaaa 8787 <210> 265 <211> 3537 <212> DNA <213> Homo sapiens <400> 265 ggcccctcga gcctcgaacc ggaacctcca aatccgagac gctctgctta tgaggacctc 60 gaaatatgcc ggccagtgaa aaaatcttgt ggctttgagg gcttttggtt ggccaggggc 120 agtaaaaatc tcggagagct gacaccaagt cctcccctgc cacgtagcag tggtaaagtc 180 cgaagctcaa attccgagaa ttgagctctg ttgattctta gaactggggt tcttagaagt 240 ggtgatgcaa gaagtttcta ggaaaggccg gacaccaggt tttgagcaaa attttggact 300 gtgaagcaag gcattggtga agacaaaatg gcctcgccgg ctgacagctg tatccagttc 360 acccgccatg ccagtgatgt tcttctcaac cttaatcgtc tccggagtcg agacatcttg 420 actgatgttg tcattgttgt gagccgtgag cagtttagag cccataaaac ggtcctcatg 480 gcctgcagtg gcctgttcta tagcatcttt acagaccagt tgaaatgcaa ccttagtgtg 540 atcaatctag atcctgagat caaccctgag ggattctgca tcctcctgga cttcatgtac 600 acatctcggc tcaatttgcg ggagggcaac atcatggctg tgatggccac ggctatgtac 660 ctgcagatgg agcatgttgt ggacacttgc cggaagttta ttaaggccag tgaagcagag 720 atggtttctg ccatcaagcc tcctcgtgaa gagttcctca acagccggat gctgatgccc 780 caagacatca tggcctatcg gggtcgtgag gtggtggaga acaacctgcc actgaggagc 840 gcccctgggt gtgagagcag agcctttgcc cccagcctgt acagtggcct gtccacaccg 900 ccagcctctt attccatgta cagccacctc cctgtcagca gcctcctctt ctccgatgag 960 gagtttcggg atgtccggat gcctgtggcc aaccccttcc ccaaggagcg ggcactccca 1020 tgtgatagtg ccaggccagt ccctggtgag tacagccggc cgactttgga ggtgtccccc 1080 aatgtgtgcc acagcaatat ctattcaccc aaggaaacaa tcccagaaga ggcacgaagt 1140 gatatgcact acagtgtggc tgagggcctc aaacctgctg ccccctcagc ccgaaatgcc 1200 ccctacttcc cttgtgacaa ggccagcaaa gaagaagaga gaccctcctc ggaagatgag 1260 attgccctgc atttcgagcc ccccaatgca cccctgaacc ggaagggtct ggttagtcca 1320 cagagccccc agaaatctga ctgccagccc aactcgccca cagagtcctg cagcagtaag 1380 aatgcctgca tcctccaggc ttctggctcc cctccagcca agagccccac tgaccccaaa 1440 gcctgcaact ggaagaaata caagttcatc gtgctcaaca gcctcaacca gaatgccaaa 1500 ccagaggggc ctgagcaggc tgagctgggc cgcctttccc cacgagccta cacggcccca 1560 cctgcctgcc agccacccat ggagcctgag aaccttgacc tccagtcccc aaccaagctg 1620 agtgccagcg gggaggactc caccatccca caagccagcc ggctcaataa catcgttaac 1680 aggtccatga cgggctctcc ccgcagcagc agcgagagcc actcaccact ctacatgcac 1740 cccccgaagt gcacgtcctg cggctctcag tccccacagc atgcagagat gtgcctccac 1800 accgctggcc ccacgttccc tgaggagatg ggagagaccc agtctgagta ctcagattct 1860 agctgtgaga acggggcctt cttctgcaat gagtgtgact gccgcttctc tgaggaggcc 1920 tcactcaaga ggcacacgct gcagacccac agtgacaaac cctacaagtg tgaccgctgc 1980 caggcctcct tccgctacaa gggcaacctc gccagccaca agaccgtcca taccggtgag 2040 aaaccctatc gttgcaacat ctgtggggcc cagttcaacc ggccagccaa cctgaaaacc 2100 cacactcgaa ttcactctgg agagaagccc tacaaatgcg aaacctgcgg agccagattt 2160 gtacaggtgg cccacctccg tgcccatgtg cttatccaca ctggtgagaa gccctatccc 2220 tgtgaaatct gtggcacccg tttccggcac cttcagactc tgaagagcca cctgcgaatc 2280 cacacaggag agaaacctta ccattgtgag aagtgtaacc tgcatttccg tcacaaaagc 2340 cagctgcgac ttcacttgcg ccagaagcat ggcgccatca ccaacaccaa ggtgcaatac 2400 cgcgtgtcag ccactgacct gcctccggag ctccccaaag cctgctgaag catggagtgt 2460 tgatgctttc gtctccagcc ccttctcaga atctacccaa aggatactgt aacactttac 2520 aatgttcatc ccatgatgta gtgcctcttt catccactag tgcaaatcat agctgggggt 2580 tgggggtggt gggggtcggg gcctggggga ctgggagccg cagcagctcc ccctccccca 2640 ctgccataaa acattaagaa aatcatattg cttcttctcc tatgtgtaag gtgaaccatg 2700 tcagcaaaaa gcaaaatcat tttatatgtc aaagcagggg agtatgcaaa agttctgact 2760 tgactttagt ctgcaaaatg aggaatgtat atgttttgtg ggaacagatg tttcttttgt 2820 atgtaaatgt gcattctttt aaaagacaag acttcagtat gttgtcaaag agagggcttt 2880 aattttttta accaaaggtg aaggaatata tggcagagtt gtaaatatat aaatatatat 2940 atatataaaa taaatatata taaacctaac aaagatatat taaaaatata aaactgcgtt 3000 aaaggctcga ttttgtatct gcaggcagac acggatctga gaatctttat tgagaaagag 3060 cacttaagag aatattttaa gtattgcatc tgtataagta agaaaatatt ttgtctaaaa 3120 tgcctcagtg tatttgtatt tttttgcaag tgaaggttta caatttacaa agtgtgtatt 3180 aaaaaaaaca aaaagaacaa aaaaatctgc agaaggaaaa atgtgtaatt ttgttctagt 3240 tttcagtttg tatatacccg tacaacgtgt cctcacggtg ccttttttca cggaagtttt 3300 caatgatggg cgagcgtgca ccatcccttt ttgaagtgta ggcagacaca gggacttgaa 3360 gttgttacta actaaactct ctttgggaat gtttgtctca tcccattctg cgtcatgctt 3420 gtgttataac tactccggag acagggtttg gctgtgtcta aactgcatta ccgcgttgta 3480 aaatatagct gtacaaatat aagaataaaa tgttgaaaag tcaaactgga aaaaaaa 3537 <210> 266 <211> 2613 <212> DNA <213> Homo sapiens <400> 266 tcccccctct taaaacacga tgcctcccag gatgctagtg gcaccactgc cactgcattt 60 cctgttggca gcagtgagca gtgaaaaccg aagcggcaga aggcagtggc agcaggcagt 120 ggcagcaggc agtggcccag gcagaaatag ctcccgcgcg attcactgga gccttccccg 180 ggccctggtc ccggctaccg ggactcgcgc gtccggatct caaaagcggc agaggccacc 240 gaagggacag gaagcacttt ggtccagacc acactcccgg cacagtgcgg aaagagccgg 300 cgggagccac tctgatcccg gacgcctcag cgcccccttg ggcttgggct tgccctcggg 360 ccggggaagg ctgaccgcga tgccaggacg cgctcccctc cgcaccgtcc cgggcgccct 420 gggtgcctgg ctgctgggcg gcctctgggc ctggaccctg tgcggcctgt gcagcctggg 480 ggcggtggga gccccgcgcc cgtgccaggc gccgcagcag tgggaggggc gccaggttat 540 gtaccagcaa agtagcgggc gcaacagccg cgccctgctc tcctacgacg ggctcaacca 600 gcgcgtgcgg gtgctggacg agaggaaggc gctgatcccc tgcaagagat tatttgaata 660 tattttgctg tataaggatg gagtgatgtt tcagattgac caagccacca agcagtgctc 720 aaagatgacc ctgacacagc cctgggatcc tcttgacatt cctcaaaact ccacctttga 780 agaccagtac tccatcgggg ggcctcagga gcagatcacc gtccaggagt ggtcggacag 840 aaagtcagct agatcctatg aaacctggat tggcatctat acagtcaagg attgctatcc 900 tgtccaggaa acctttacca taaactacag tgtgatattg tctacgcggt tttttgacat 960 ccagctgggt attaaagacc cctcggtgtt tacccctcca agcacgtgcc agatggccca 1020 actggagaag atgagcgaag actgctcctg gtgagcctgt gcatagggaa gcggcagcat 1080 cggatgtcag ccccctgcgg ccccagctgg agatggatat gagactagtc aagatgtgaa 1140 tgctaattgg agagaaatat aattttagga agatgcacat tgatgtgggg ttttgatgtg 1200 tctgattttg actactcaag ctctgtttac agaagaaaat tgaatggcga gggtgtggcc 1260 atatgaactg actagatggc taatatggac actttgggta tttctaatgc ctgttcaggg 1320 ctggttttct gcatgcacgg gtatacacat aatgcagtgc catgcacata gggaagggtc 1380 agtaagagaa gtttgccttg gcagcaagta tttattgttg acattattca gaattagtga 1440 taataaaaag cagagtgatt ttggtcaatt ttattattaa ttcttaaatt ccctgcagag 1500 aatgccccct ttattgctgc accagggttg gcattgctcc cactgagccc tactccaccc 1560 tgtccctgca ctcccttggt tgccaaaaaa atgataactt aaatcccttc cagacttaag 1620 aattttatgg catggcccaa ttgatataaa catttagaag gaaatgaaaa gctaaaatag 1680 gaagtaatta ttcctctaaa gaaacatttt gagcaaggca gtttagagaa tcctaatgtc 1740 tacactggca tagcacgagc catgtaagct tctttttttt ctatgcaaga gtattgatgt 1800 atgtgctgaa tcttcacaga cttgtcaata cacaggcagt attctaaaat agcactgaac 1860 agggagtcag gagactattg tctcctaaac ccaggactag agttccctcg tactgtcact 1920 cctttggtca ttaaatgcac tgggcttgcc cgcactttgg ccttcctaga acactgcttc 1980 ataacctctc tgtctgactt ctgcatctcc ttccaggtca gctcattcac aagagttgct 2040 cccaagcctg gatgagttgc accttgcatc ttgagcatgc atttctcaca ataattatta 2100 agctgtgtga taatttctgc tttcaggaca ctcatccatt atcttggctg tgagctcctt 2160 gggtacgggt accttgtatg tttactttta tatccctagc acaaagcaag tgcctggcac 2220 atagtcagtg ccctaagtat tcgtagagtg aagaatgcca gcctctcttg tccctggttt 2280 ccttatgtgt tgaatgtggt tgagtttgtc cattgctagg gagagacttc cagtaataaa 2340 atttactatt ctagatgctt ctactgttat gttttatctg cccatttatc tttcttagtt 2400 accaggagaa atgtgtgaca cctatattat aatgaaaaca atctcattac ttatagttta 2460 tctatattaa acaaatttaa ttgcatttta aagcattctt tgatactgtt gcttttgcaa 2520 taaatatgga taatcttggt tataagggag ttaaaacaat gctgtaataa ataaagtgct 2580 tcatgtgatc aaaatcaaaa aaaaaaaaaa aaa 2613 <210> 267 <211> 1890 <212> DNA <213> Homo sapiens <400> 267 ctagagaggc cgccaggaga cccggcgctt tcttccttct gcagctgagg ctgcggcggg 60 gccggggctg gggtcggggc caggaggaat tttgttgtca gagaataaaa ggaggttgtc 120 cataattgac tttaagcagc aatcagtaaa acattgagct cttcagctcc gcctttcttg 180 ctctgaaaat tggaaaacca agaaggtttt gatgttttgt gtgacgccac ctgaattaga 240 aaccaagatg aacataacca aaggtggtct ggtgttgttt tcagcaaact cgaattcatc 300 atgtatggag ctatcaaaga aaattgcaga gcggctaggg gtggagatgg gcaaagtgca 360 ggtttaccag gaacctaaca gagaaacaag agtacaaatt caagagtctg tgaggggaaa 420 agatgttttc atcatccaaa ctgtttcgaa ggacgtgaac accaccatca tggagctcct 480 gatcatggtg tatgcatgta agacctcttg tgccaagagc atcattggcg tgatacccta 540 ctttccttac agcaagcagt gcaagatgag aaaaagaggc tccattgtct ctaaattgct 600 ggcttccatg atgtgcaaag ctggtctaac tcatcttatt actatggatt tacaccagaa 660 ggaaattcag ggcttcttca atattcctgt tgacaattta agagcatctc ccttcttatt 720 acagtatatt caagaagaga tcccagatta caggaatgca gtaatcgtgg ccaagtctcc 780 agcctcggcg aagagggcac agtcttttgc tgagcgcctg cgcctgggaa ttgcagtgat 840 tcatggagag gcgcaggatg ccgagtcgga cttggtggat ggacggcatt ccccacccat 900 ggtcagaagt gtggctgcca tccaccccag cctggagatc cccatgctga ttcctaaaga 960 aaagccccca atcacggttg tgggtgatgt tggaggaagg attgccatca tcgtggatga 1020 catcattgat gatgttgaca gctttcttgc tgcagcagag accctgaagg aaagaggtgc 1080 atataagatc tttgtgatgg caactcatgg cttgttgtct tctgacgccc cccggcggat 1140 tgaagagtct gccattgatg aggtggtggt caccaataca attccacatg aagtccagaa 1200 gctccagtgc cccaagatta aaactgtgga tatcagcatg atcctttcag aggcgatccg 1260 tcggatccac aatggggagt ccatgtccta ccttttcaga aacataggct tagatgactg 1320 agttttcctt taggaaaact cccgagggcc aaactggaaa cataagagtg actgctcggt 1380 gggatggatt tcacaggaac cgtcatgctt gttcctccct ctcccctgta acctcacttc 1440 ttattgattc ctaagaagat agaccaactt tttatgtcgg tttgggtgtt tgtgagtttg 1500 gggagcaatt tttataaaag aaaaacttta ttctcctctt ttgaaaaggt aagacctcgt 1560 tttagttgta actgtttaaa aaataacact tggaataaga tttgtaagct cacaaagcct 1620 tcttccaaag ttgcttgagc caagtgctta aaaagttaat aaaataaaat gatctgtatg 1680 atacctgcaa ttgaaaagcc gaaaagatta tactgtcaag tccagtaaat gacattttta 1740 gagatgcttt tgtagacaag catatggaat atgtgattgt atttattttc tgcaactaaa 1800 aaaggaataa aaacttgtgt ttgtgtgttt ttctaaaact ttgtgttttg gcaatcgttt 1860 tataactaaa ataaaatgaa agctaaatct 1890 <210> 268 <211> 11242 <212> DNA <213> Homo sapiens <400> 268 tttttttttt ttttttttga gaaaggggaa tttcatccca aataaaagga atgaagtctg 60 gctccggagg agggtccccg acctcgctgt gggggctcct gtttctctcc gccgcgctct 120 cgctctggcc gacgagtgga gaaatctgcg ggccaggcat cgacatccgc aacgactatc 180 agcagctgaa gcgcctggag aactgcacgg tgatcgaggg ctacctccac atcctgctca 240 tctccaaggc cgaggactac cgcagctacc gcttccccaa gctcacggtc attaccgagt 300 acttgctgct gttccgagtg gctggcctcg agagcctcgg agacctcttc cccaacctca 360 cggtcatccg cggctggaaa ctcttctaca actacgccct ggtcatcttc gagatgacca 420 atctcaagga tattgggctt tacaacctga ggaacattac tcggggggcc atcaggattg 480 agaaaaatgc tgacctctgt tacctctcca ctgtggactg gtccctgatc ctggatgcgg 540 tgtccaataa ctacattgtg gggaataagc ccccaaagga atgtggggac ctgtgtccag 600 ggaccatgga ggagaagccg atgtgtgaga agaccaccat caacaatgag tacaactacc 660 gctgctggac cacaaaccgc tgccagaaaa tgtgcccaag cacgtgtggg aagcgggcgt 720 gcaccgagaa caatgagtgc tgccaccccg agtgcctggg cagctgcagc gcgcctgaca 780 acgacacggc ctgtgtagct tgccgccact actactatgc cggtgtctgt gtgcctgcct 840 gcccgcccaa cacctacagg tttgagggct ggcgctgtgt ggaccgtgac ttctgcgcca 900 acatcctcag cgccgagagc agcgactccg aggggtttgt gatccacgac ggcgagtgca 960 tgcaggagtg cccctcgggc ttcatccgca acggcagcca gagcatgtac tgcatccctt 1020 gtgaaggtcc ttgcccgaag gtctgtgagg aagaaaagaa aacaaagacc attgattctg 1080 ttacttctgc tcagatgctc caaggatgca ccatcttcaa gggcaatttg ctcattaaca 1140 tccgacgggg gaataacatt gcttcagagc tggagaactt catggggctc atcgaggtgg 1200 tgacgggcta cgtgaagatc cgccattctc atgccttggt ctccttgtcc ttcctaaaaa 1260 accttcgcct catcctagga gaggagcagc tagaagggaa ttactccttc tacgtcctcg 1320 acaaccagaa cttgcagcaa ctgtgggact gggaccaccg caacctgacc atcaaagcag 1380 ggaaaatgta ctttgctttc aatcccaaat tatgtgtttc cgaaatttac cgcatggagg 1440 aagtgacggg gactaaaggg cgccaaagca aaggggacat aaacaccagg aacaacgggg 1500 agagagcctc ctgtgaaagt gacgtcctgc atttcacctc caccaccacg tcgaagaatc 1560 gcatcatcat aacctggcac cggtaccggc cccctgacta cagggatctc atcagcttca 1620 ccgtttacta caaggaagca ccctttaaga atgtcacaga gtatgatggg caggatgcct 1680 gcggctccaa cagctggaac atggtggacg tggacctccc gcccaacaag gacgtggagc 1740 ccggcatctt actacatggg ctgaagccct ggactcagta cgccgtttac gtcaaggctg 1800 tgaccctcac catggtggag aacgaccata tccgtggggc caagagtgag atcttgtaca 1860 ttcgcaccaa tgcttcagtt ccttccattc ccttggacgt tctttcagca tcgaactcct 1920 cttctcagtt aatcgtgaag tggaaccctc cctctctgcc caacggcaac ctgagttact 1980 acattgtgcg ctggcagcgg cagcctcagg acggctacct ttaccggcac aattactgct 2040 ccaaagacaa aatccccatc aggaagtatg ccgacggcac catcgacatt gaggaggtca 2100 cagagaaccc caagactgag gtgtgtggtg gggagaaagg gccttgctgc gcctgcccca 2160 aaactgaagc cgagaagcag gccgagaagg aggaggctga ataccgcaaa gtctttgaga 2220 atttcctgca caactccatc ttcgtgccca gacctgaaag gaagcggaga gatgtcatgc 2280 aagtggccaa caccaccatg tccagccgaa gcaggaacac cacggccgca gacacctaca 2340 acatcaccga cccggaagag ctggagacag agtacccttt ctttgagagc agagtggata 2400 acaaggagag aactgtcatt tctaaccttc ggcctttcac attgtaccgc atcgatatcc 2460 acagctgcaa ccacgaggct gagaagctgg gctgcagcgc ctccaacttc gtctttgcaa 2520 ggactatgcc cgcagaagga gcagatgaca ttcctgggcc agtgacctgg gagccaaggc 2580 ctgaaaactc catcttttta aagtggccgg aacctgagaa tcccaatgga ttgattctaa 2640 tgtatgaaat aaaatacgga tcacaagttg aggatcagcg agaatgtgtg tccagacagg 2700 aatacaggaa gtatggaggg gccaagctaa accggctaaa cccggggaac tacacagccc 2760 ggattcaggc cacatctctc tctgggaatg ggtcgtggac agatcctgtg ttcttctatg 2820 tccaggccaa aacaggatat gaaaacttca tccatctgat catcgctctg cccgtcgctg 2880 tcctgttgat cgtgggaggg ttggtgatta tgctgtacgt cttccataga aagagaaata 2940 acagcaggct ggggaatgga gtgctgtatg cctctgtgaa cccggagtac ttcagcgctg 3000 ctgatgtgta cgttcctgat gagtgggagg tggctcggga gaagatcacc atgagccggg 3060 aacttgggca ggggtcgttt gggatggtct atgaaggagt tgccaagggt gtggtgaaag 3120 atgaacctga aaccagagtg gccattaaaa cagtgaacga ggccgcaagc atgcgtgaga 3180 ggattgagtt tctcaacgaa gcttctgtga tgaaggagtt caattgtcac catgtggtgc 3240 gattgctggg tgtggtgtcc caaggccagc caacactggt catcatggaa ctgatgacac 3300 ggggcgatct caaaagttat ctccggtctc tgaggccaga aatggagaat aatccagtcc 3360 tagcacctcc aagcctgagc aagatgattc agatggccgg agagattgca gacggcatgg 3420 catacctcaa cgccaataag ttcgtccaca gagaccttgc tgcccggaat tgcatggtag 3480 ccgaagattt cacagtcaaa atcggagatt ttggtatgac gcgagatatc tatgagacag 3540 actattaccg gaaaggaggg aaagggctgc tgcccgtgcg ctggatgtct cctgagtccc 3600 tcaaggatgg agtcttcacc acttactcgg acgtctggtc cttcggggtc gtcctctggg 3660 agatcgccac actggccgag cagccctacc agggcttgtc caacgagcaa gtccttcgct 3720 tcgtcatgga gggcggcctt ctggacaagc cagacaactg tcctgacatg ctgtttgaac 3780 tgatgcgcat gtgctggcag tataacccca agatgaggcc ttccttcctg gagatcatca 3840 gcagcatcaa agaggagatg gagcctggct tccgggaggt ctccttctac tacagcgagg 3900 agaacaagct gcccgagccg gaggagctgg acctggagcc agagaacatg gagagcgtcc 3960 ccctggaccc ctcggcctcc tcgtcctccc tgccactgcc cgacagacac tcaggacaca 4020 aggccgagaa cggccccggc cctggggtgc tggtcctccg cgccagcttc gacgagagac 4080 agccttacgc ccacatgaac gggggccgca agaacgagcg ggccttgccg ctgccccagt 4140 cttcgacctg ctgatccttg gatcctgaat ctgtgcaaac agtaacgtgt gcgcacgcgc 4200 agcggggtgg ggggggagag agagttttaa caatccattc acaagcctcc tgtacctcag 4260 tggatcttca gaactgccct tgctgcccgc gggagacagc ttctctgcag taaaacacat 4320 ttgggatgtt ccttttttca atatgcaagc agctttttat tccctgccca aacccttaac 4380 tgacatgggc ctttaagaac cttaatgaca acacttaata gcaacagagc acttgagaac 4440 cagtctcctc actctgtccc tgtccttccc tgttctccct ttctctctcc tctctgcttc 4500 ataacggaaa aataattgcc acaagtccag ctgggaagcc ctttttatca gtttgaggaa 4560 gtggctgtcc ctgtggcccc atccaaccac tgtacacacc cgcctgacac cgtgggtcat 4620 tacaaaaaaa cacgtggaga tggaaatttt tacctttatc tttcaccttt ctagggacat 4680 gaaatttaca aagggccatc gttcatccaa ggctgttacc attttaacgc tgcctaattt 4740 tgccaaaatc ctgaactttc tccctcatcg gcccggcgct gattcctcgt gtccggaggc 4800 atgggtgagc atggcagctg gttgctccat ttgagagaca cgctggcgac acactccgtc 4860 catccgactg cccctgctgt gctgctcaag gccacaggca cacaggtctc attgcttctg 4920 actagattat tatttggggg aactggacac aataggtctt tctctcagtg aaggtgggga 4980 gaagctgaac cggcttccct gccctgcctc cccagccccc tgcccaaccc ccaagaatct 5040 ggtggccatg ggccccgaag cagcctggcg gacaggcttg gagtcaaggg gccccatgcc 5100 tgcttctctc ccagccccag ctcccccgcc cgcccccaag gacacagatg ggaaggggtt 5160 tccagggact cagccccact gttgatgcag gtttgcaagg aaagaaattc aaacaccaca 5220 acagcagtaa gaagaaaagc agtcaatgga ttcaagcatt ctaagctttg ttgacatttt 5280 ctctgttcct aggacttctt catgggtctt acagttctat gttagaccat gaaacatttg 5340 catacacatc gtctttaatg tcacttttat aactttttta cggttcagat attcatctat 5400 acgtctgtac agaaaaaaaa aagctgctat tttttttgtt cttgatcttt gtggatttaa 5460 tctatgaaaa ccttcaggtc caccctctcc cctttctgct cactccaaga aacttcttat 5520 gctttgtact agagtgcgtg actttcttcc tcttttcccg gtaatggata cttctatcac 5580 ataatttgcc atgaactgtt ggatgccttt ttataaatac atcccccatc cctgctccca 5640 cctgcccctt tagttgtttt ctaacccgta ggctctctgg gcacgaggca gaaagcaggc 5700 cgggcaccca tcctgagagg gccgcgctcc tctccccagc ctgccctcac agcattggag 5760 cctgttacag tgcaagacat gatacaaact caggtcagaa aaacaaaggt taaatatttc 5820 acacgtcttt gttcagtgtt tccactcacc gtggttgaga agcctcaccc tctctttccc 5880 ttgcctttgc ttaggttgtg acacacatat atatatattt ttttaattct tgggtacaac 5940 agcagtgtta accgcagaca ctaggcattt ggattactat ttttcttaat ggctatttaa 6000 tccttccatc ccacgaaaaa cagctgctga gtccaaggga gcagcagagc gtggtccggc 6060 agggcctgtt gtggccctcg ccacccccct caccggaccg actgacctgt ctttggaacc 6120 agaacatccc aagggaactc cttcgcactg gcgttgagtg ggaccccggg atccaggctg 6180 gcccagggcg gcaccctcag ggctgtgccc gctggagtgc taggtggagg cagcacagac 6240 gccacggtgg cccaagagcc cctttgcttc ttgctggggg accagggctg tggtgctggc 6300 ccactttccc tcggccagga atccaggtcc ttggggccca ggggtcttgt cttgtttcat 6360 ttttagcact tctcaccaga gagatgacag cacaagagtt gcttctggga tagaaatgtt 6420 taggagtaag aacaaagctg ggatacggtg attgctagtt gtgactgaag attcaacaca 6480 gaaaagaaag tttatacggc ttttttgctg gtcagcagtt tgtcccactg ctttctctag 6540 tctctatccc atagcgtgtt ccctttaaaa aaaaaaaaaa ggtattatat gtaggagttt 6600 tcttttaatt tattttgtga taaattacca gtttcaatca ctgtagaaaa gccccattat 6660 gaatttaaat ttcaaggaaa gggtgtgtgt gtgtgtatgt gtggggtgtg tgtgtgtgag 6720 agtgatggga cagttcttga ttttttgggt tttttttccc ccaaacattt atctacctca 6780 ctcttatttt ttatatgtgt atatagacaa aagaatacat ctcacctttc tcagcacctg 6840 acaataggcc gttgatactg gtaacctcat ccacgccaca ggcgccacac ccaggtgatg 6900 cagggggaag ccaggctgta ttccggggtc aaagcaacac taactcacct ctctgctcat 6960 ttcagacagc ttgccttttt ctgagatgtc ctgttttgtg ttgctttttt tgttttgttt 7020 tctatcttgg tttccaccaa ggtgttagat ttctcctcct cctagccagg tggccctgtg 7080 aggccaacga gggcaccaga gcacacctgg gggagccacc aggctgtccc tggctggttg 7140 tctttggaac aaactgcttc tgtgcagatg gaatgaccaa cacatttcgt ccttaagaga 7200 gcagtggttc ctcaggttct gaggagagga aggtgtccag gcagcaccat ctctgtgcga 7260 atccccaggg taaaggcgtg gggcattggg tttgctcccc ttgctgctgc tccatccctg 7320 caggaggctc gcgctgaggc aggaccgtgc ggccatggct gctgcattca ttgagcacaa 7380 aggtgcagct gcagcagcag ctggagagca agagtcaccc agcctgtgcg ccagaatgca 7440 gaggctcctg acctcacagc cagtccctga tagaacacac gcaggagcag agtcccctcc 7500 ccctccaggc tgccctctca acttctccct cacctccttc cctaggggta gacagagatg 7560 taccaaacct tccggctgga aagcccagtg gccggcgccg aggctcgtgg cgtcacgccc 7620 cccccgccag ggctgtacct ccgtctccct ggtcctgctg ctcacaggac agacggctcg 7680 ctcccctctt ccagcagctg ctcttacagg cactgatgat ttcgctggga agtgtggcgg 7740 gcagctttgc ctaagcgtgg atggctcctc ggcaattcca gcctaagtga aggcgctcag 7800 gagcctcctg ctggaacgcg acccatctct cccaggaccc cggggatctt aaggtcattg 7860 agaaatactg ttggatcagg gttttgttct tccacactgt aggtgacccc ttggaataac 7920 ggcctctcct ctcgtgcaca tacctaccgg tttccacaac tggatttcta cagatcattc 7980 agctggttat aagggttttg tttaaactgt ccgagttact gatgtcattt tgtttttgtt 8040 ttatgtaggt agcttttaag tagaaaacac taacagtgta gtgcccatca tagcaaatgc 8100 ttcagaaaca cctcaataaa agagaaaact tggcttgtgt gatggtgcag tcactttact 8160 ggaccaaccc acccaccttg actataccaa ggcatcatct atccacagtt ctagcctaac 8220 ttcatgctga tttctctgcc tcttgatttt tctctgtgtg ttccaaataa tcttaagctg 8280 agttgtggca ttttccatgc aacctccttc tgccagcagc tcacactgct tgaagtcata 8340 tgaaccactg aggcacatca tggaattgat gtgagcatta agacgttctc ccacacagcc 8400 cttccctgag gcagcaggag ctggtgtgta ctggagacac tgttgaactt gatcaagacc 8460 cagaccaccc caggtctcct tcgtgggatg tcatgacgtt tgacatacct ttggaacgag 8520 cctcctcctt ggaagatgga agaccgtgtt cgtggccgac ctggcctctc ctggcctgtt 8580 tcttaagatg cggagtcaca tttcaatggt acgaaaagtg gcttcgtaaa atagaagagc 8640 agtcactgtg gaactaccaa atggcgagat gctcggtgca cattggggtg ctttgggata 8700 aaagatttat gagccaacta ttctctggca ccagattcta ggccagtttg ttccactgaa 8760 gcttttccca cagcagtcca cctctgcagg ctggcagccg aatggcttgc cagtggctct 8820 gtggcaagat cacactgaga tcgatgggtg agaaggctag gatgcttgtc tagtgttctt 8880 agctgtcacg ttggctcctt ccagggtggc cagacggtgt tggccactcc cttctaaaac 8940 acaggcgccc tcctggtgac agtgacccgc cgtggtatgc cttggcccat tccagcagtc 9000 ccagttatgc atttcaagtt tggggtttgt tcttttcgtt aatgttcctc tgtgttgtca 9060 gctgtcttca tttcctgggc taagcagcat tgggagatgt ggaccagaga tccactcctt 9120 aagaaccagt ggcgaaagac actttctttc ttcactctga agtagctggt ggtacaaatg 9180 agaacttcaa gagaggatgt tatttagact gaacctctgt tgccagagat gctgaagata 9240 cagaccttgg acaggtcaga gggtttcatt tttggccttc atcttagatg actggttgcg 9300 tcatttggag aagtgagtgc tccttgatgg tggaatgacc gggtggtggg tacagaacca 9360 ttgtcacagg gatcctggca cagagaagag ttacgagcag cagggtgcag ggcttggaag 9420 gaatgtgggc aaggttttga acttgattgt tcttgaagct atcagaccac atcgaggctc 9480 agcagtcatc cgtgggcatt tggtttcaac aaagaaacct aacatcctac tctggaaact 9540 gatctcggag ttaaggcgaa ttgttcaaga acacaaacta catcgcactc gtcagttgtc 9600 agttctgggg catgacttta gcgttttgtt tctgcgagaa cataacgatc actcattttt 9660 atgtcccacg tgtgtgtgtc cgcatctttc tggtcaacat tgttttaact agtcactcat 9720 tagcgttttc aatagggctc ttaagtccag tagattacgg gtagtcagtt gacgaagatc 9780 tggtttacaa gaactaatta aatgtttcat tgcatttttg taagaacaga ataattttat 9840 aaaatgtttg tagtttataa ttgccgaaaa taatttaaag acactttttt tttctctgtg 9900 tgtgcaaatg tgtgtttgtg atccattttt tttttttttt tttaggacac ctgtttacta 9960 gctagcttta caatatgcca aaaaaggatt tctccctgac cccatccgtg gttcaccctc 10020 ttttcccccc atgctttttg ccctagttta taacaaagga atgatgatga tttaaaaagt 10080 agttctgtat cttcagtatc ttggtcttcc agaaccctct ggttgggaag gggatcattt 10140 tttactggtc atttcccttt ggagtgtagc tactttaaca gatggaaaga acctcattgg 10200 ccatggaaac agccgaggtg ttggagccca gcagtgcatg gcaccgttcg gcatctggct 10260 tgattggtct ggctgccgtc attgtcagca cagtgccatg gacatgggaa gacttgactg 10320 cacagccaat ggttttcatg atgattacag catacacagt gatcacataa acgatgacag 10380 ctatggggca cacaggccat ttgcttacat gcctcgtatc atgactgatt actgctttgt 10440 tagaacacag aagagaccct attttattta aggcagaacc ccgaagatac gtatttccaa 10500 tacagaaaag aatttttaat aaaaactata acatacacaa aaattggttt taaagttgac 10560 tccacttcct ctaactccag tggattgttg gccatgtctc cccaactcca caatatctct 10620 atcatgggaa acacctgggg tttttgcgct acataggaga aagatctgga aactatttgg 10680 gttttgtttt caacttttca tttggatgtt tggcgttgca cacacacatc caccggtgga 10740 agagacgccc ggtgaaaaca cctgtctgct ttctaagcca gtgaggttga ggtgagaggt 10800 ttgccagagt ttgtctacct ctgggtatcc ctttgtctgg gataaaaaaa atcaaaccag 10860 aaggcgggat ggaatggatg caccgcaaat aatgcatttt ctgagttttc ttgttaaaaa 10920 aaaatttttt taagtaagaa aaaaaaaggt aataacatgg ccaatttgtt acataaaatg 10980 actttctgtg tataaattat tcctaaaaaa tcctgtttat ataaaaaatc agtagatgaa 11040 aaaaatttca aaatgttttt gtatattctg ttgtaagaat ttattcctgt tattgcgata 11100 tactctggat tctttacata atggaaaaaa gaaactgtct attttgaatg gctgaagcta 11160 aggcaacgtt agtttctctt actctgcttt tttctagtaa agtactacat ggtttaagtt 11220 aaataaaata attctgtatg ca 11242 <210> 269 <211> 2718 <212> DNA <213> Homo sapiens <400> 269 cagggtaacg ctgtcttgtg gacccgcact tcccacccga gacctctcac tgagcccgag 60 ccgcgcgcga catgagccac gggaagggaa ccgacatgct cccggagatc gccgccgccg 120 tgggcttcct ctccagcctc ctgaggaccc ggggctgcgt gagcgagcag aggcttaagg 180 tcttcagcgg ggcgctccag gaggcactca cagagcacta caaacaccac tggtttcccg 240 aaaagccgtc caagggctcc ggctaccgct gcattcgcat caaccacaag atggacccca 300 tcatcagcag ggtggccagc cagatcggac tcagccagcc ccagctgcac cagctgctgc 360 ccagcgagct gaccctgtgg gtggacccct atgaggtgtc ctaccgcatt ggggaggacg 420 gctccatctg cgtcttgtac gaggaggccc cactggccgc ctcctgtggg ctcctcacct 480 gcaagaacca agtgctgctg ggccggagca gcccctccaa gaactacgtg atggcagtct 540 ccagctaggc ccttccgccc ccgccctggg cgccgccgtg ctcatgctgc cgtgacaaca 600 ggccaccaca tacctcaacc tggggaactg tatttttaaa tgaagagcta tttatatata 660 ttattttttt ttaagaaagg aggaaaagaa accaaaagtt ttttttaaga aaaaaaatcc 720 ttcaagggag ctgcttggaa gtggcctccc caggtgcctt tggagagaac tgttgcgtgc 780 ttgagtctgt gagccagtgt ctgcctatag gagggggagc tgttaggggg tagacctagc 840 caaggagaag tgggagacgt ttggctagca ccccaggaag atgtgagagg gagcaagcaa 900 ggttagcaac tgtgaacaga gaggtcggga tttgccctgg gggaggaaga gaggccaagt 960 tcagagctct ctgtctcccc cagccagaca cctgcatccc tggctcctct attactcagg 1020 ggcattcatg cctggactta aacaatacta tgttatcttt tcttttattt ttctaatgag 1080 gtcctgggca gagagtgaaa aggcctctcc tgattcctac tgtcctaagc tgcttttctt 1140 gaaatcatga cttgtttcta attctaccct caggggcctg tagatgttgc tttccagcca 1200 ggaatctaaa gctttgggtt ttctgagggg ggggaggagg gaactggagg ttattggggt 1260 taggatggaa gggaactctg cacaaaacct ttgctttgct agtgctgctt tgtgtgtatg 1320 tgtggcaaat aatttggggg tgatttgcaa tgaaattttg ggacccaaag agtatccact 1380 ggggatgttt tttggccaaa actcttcctt ttggaaccac atgaaagtct tgatgctgct 1440 gccatgatcc ctttgagagg tggctcaaaa gctacaggga actccaggtc ctttattact 1500 gccttctttt caaaagcaca actctcctct aaccctcccc tcccccttcc cttctggtcg 1560 ggtcatagag ctaccgtatt ttctaggaca agagttctca gtcactgtgc aatatgcccc 1620 ctgggtccca ggagggtctg gaggaaaact ggctatcaga acctcctgat gccctggtgg 1680 gcttagggaa ccatctctcc tgctctcctt gggatgatgg ctggctagtc agccttgcat 1740 gtattccttg gctgaatggg agagtgcccc atgttctgca agactacttg gtattcttgt 1800 agggccgaca ctaaataaaa gccaaacctt gggcactgtt ttttctccct ggtgctcaga 1860 gcacctgtgg gaaaggttgc tgtctgtctc agtacaatcc aaatttgtcg tagacttgtg 1920 caatatatac tgttgtgggt tggagaaaag tggaaagcta cactgggaag aaactccctt 1980 ccttcaattt ctcagtgaca ttgatgaggg gtcctcaaaa gacctcgagt ttcccaaacc 2040 gaatcacctt aagaaggaca gggctagggc atttggccag gatggccacc ctcctgctgt 2100 tgccccttag tgaggaatct tcaccccact tcctctaccc ccaggttctc ctccccacag 2160 ccagtcccct ttcctggatt tctaaactgc tcaattttga ctcaaaggtg ctatttacca 2220 aacactctcc ctacccattc ctgccagctc tgcctccttt tcaactctcc acattttgta 2280 ttgccttccc agacctgctt ccagtcttta ttgctttaaa gttcactttg ggcccacaga 2340 cccaagagct aattttctgg tttgtgggtt gaaacaaagc tgtgaatcac tgcaggctgt 2400 gttcttgcat cttgtctgca aacaggtccc tgccttttta gaagcagcct catggtctca 2460 tgcttaatct tgtctctctt ctcttcttta tgatgttcac tttaaaaaca acaaaacccc 2520 tgagctggac tgttgagcag gcctgtctct cctattaagt aaaaataaat agtagtagta 2580 tgtttgtaag ctattctgac agaaaagaca aaggttacta attgtatgat agtgttttta 2640 tatggaagaa tgtacagctt atggacaaat gtacaccttt ttgttacttt aataaaaatg 2700 tagtaggata aaaaaaaa 2718 <210> 270 <211> 2304 <212> DNA <213> Homo sapiens <400> 270 gaattcgtcc aaactgagga tcacaagtct ccacattctg agtaggagga tgagggtctg 60 agttaggatt tgggtcctgc agggcttgct aaggaatccc ctgatggcct aggattccac 120 gcagagcaca tctggtgtga gagagctcgc tgcaagggtg aaggctccgc cctatcagat 180 agacaaccag gccaccaaga ggcccagccc tccaaaccct ggatttgcaa catcctcaaa 240 gaacagcaac gggccttgag cagaattgag aaggaaatac ccccacctgc cctcagccgt 300 taagtgggct ttgctattca caagggcctc tgggtgtcct ggcagagagg ggagatggca 360 caggcaccag gtgctagggt gccagggcct cccgagaagg aacaggtgca aagcaggcaa 420 ttagcccaga aggtatccgt ggggcaggca gcctagatct gatgggggaa gccaccagga 480 ttacatcatc tgctgtaaca actgctctga aaagaagata tttttcaacc tgaacttgca 540 gtagctagtg gagaggcagg aaaaaggaaa tgaaacagag acagagggaa gcctgagcca 600 aaatagacct tcccgagaga ggaggaagcc cggagagaga cgcacggtcc cctccccgcc 660 cctaggccgc cgccccctct ctgccctcgg cggcgagcag ggcgccgcga cccggggccg 720 gaaaggtgcc aggggctccg ggcggccggg cgggcgcaca ccatccccgc gggcggcgcg 780 gagccggcga cagcgcgcga gagggaccgg gcggtggcgg cggcgggacc gggatggaag 840 ggagcgcggt gactgtcctt gagcgcggag gggcgagctc gccggcggag gccgagcaag 900 cggaggcagg agcggcggcg acggcggcgg cggcggcggc gcccgagcac ccgagggggt 960 ccgagccccg gcagccggcc agccccgcgc cacaaaggga gcgcccccgc cgcccggcac 1020 cccgcctccc tccccaatgt cctcggccat cgaaaggaag agcctggacc cttcagagga 1080 accagtggat gaggtgctgc agatcccccc atccctgctg acatgcggcg gctgccagca 1140 gaacatcggg gaccgctact tcctgaaggc catcgaccag tactggcacg aggactgcct 1200 gagctgcgac ctctgtggct gccggctggg tgaggtgggg cggcgcctct actacaaact 1260 gggccggaag ctctgccgga gagactatct caggcttttt gggcaagacg gtctctgcgc 1320 atcctgtgac aagcggattc gtgcctatga gatgacaatg cgggtgaaag acaaagtgta 1380 tcacctggaa tgtttcaagt gcgccgcctg tcagaagcat ttctgtgtag gtgacagata 1440 cctcctcatc aactctgaca tagtgtgcga acaggacatc tacgagtgga ctaagatcaa 1500 tgggatgata taggcccgag tccccgggca tctttgggga ggtgttcact gaagacgccg 1560 tctccatggc atcttcgtct tcactcttag gcactttggg ggtttgaggg tggggtaagg 1620 gatttcttag gggatggtag acctttattg ggtatcaaga catagcatcc aagtggcata 1680 attcaggggc tgacacttca aggtgacaga aggaccagcc cttgagggag aacttatggc 1740 cacagcccat ccatagtaac tgacatgatt agcagaagaa aggaacattt aggggcaagc 1800 aggcgctgtg ctatcatgat ggaatttcat atctacagat agagagttgt tgtgtacaga 1860 cttgttgtga ctttgacgct tgcgaactag agatgtgcaa ttgatttctt ttcttcctgg 1920 ctttttaact cccctgtttc aatcactgtc ctccacacaa gggaaggaca gaaaggagag 1980 tggccattct ttttttcttg gcccccttcc caaggcctta agctttggac ccaagggaaa 2040 actgcatgga gacgcatttc ggttgagaat ggaaaccaca acttttaacc aaacaattat 2100 ttaaagcaat gctgatgaat cactgttttt agacaccttc attttgaggg gaggagttcc 2160 acagattgtt tctatacaaa tataaatctt aaaaagttgt tcaactattt tattatccta 2220 gattatatca aagtatttgt cgtgtgtaga aaaaaaaaac agctctgcag gcttaataaa 2280 aatgacagac tgaaaaaaaa aaaa 2304 <210> 271 <211> 1554 <212> DNA <213> Homo sapiens <400> 271 gcttctcctt tttgtgttcc ggccgatccc acctctcctc gaccctggac gtctaccttc 60 cggaggccca catcttgccc actccgcgcg cggggctagc gcgggtttca gcgacgggag 120 ccctcaaggg acatggcaac tacagcggcg ccggcgggcg gcgcccgaaa tggagctggc 180 ccggaatggg gagggttcga agaaaacatc cagggcggag gctcagctgt gattgacatg 240 gagaacatgg atgatacctc aggctctagc ttcgaggata tgggtgagct gcatcagcgc 300 ctgcgcgagg aagaagtaga cgctgatgca gctgatgcag ctgctgctga agaggaggat 360 ggagagttcc tgggcatgaa gggctttaag ggacagctga gccggcaggt ggcagatcag 420 atgtggcagg ctgggaaaag acaagcctcc agggccttca gcttgtacgc caacatcgac 480 atcctcagac cctactttga tgtggagcct gctcaggtgc gaagcaggct cctggagtcc 540 atgatcccta tcaagatggt caacttcccc cagaaaattg caggtgaact ctatggacct 600 ctcatgctgg tcttcactct ggttgctatc ctactccatg ggatgaagac gtctgacact 660 attatccggg agggcaccct gatgggcaca gccattggca cctgcttcgg ctactggctg 720 ggagtctcat ccttcattta cttccttgcc tacctgtgca acgcccagat caccatgctg 780 cagatgttgg cactgctggg ctatggcctc tttgggcatt gcattgtcct gttcatcacc 840 tataatatcc acctccacgc cctcttctac ctcttctggc tgttggtggg tggactgtcc 900 acactgcgca tggtagcagt gttggtgtct cggaccgtgg gccccacaca gcggctgctc 960 ctctgtggca ccctggctgc cctacacatg ctcttcctgc tctatctgca ttttgcctac 1020 cacaaagtgg tagaggggat cctggacaca ctggagggcc ccaacatccc gcccatccag 1080 agggtcccca gagacatccc tgccatgctc cctgctgctc ggcttcccac caccgtcctc 1140 aacgccacag ccaaagctgt tgcggtgacc ctgcagtcac actgacccca cctgaaattc 1200 ttggccagtc ctctttcccg cagctgcaga gaggaggaag actattaaag gacagtcctg 1260 atgacatgtt tcgtagatgg ggtttgcagc tgccactgag ctgtagctgc gtaagtacct 1320 ccttgatgcc tgtcggcact tctgaaaggc acaaggccaa gaactcctgg ccaggactgc 1380 aaggctctgc agccaatgca gaaaatgggt cagctccttt gagaacccct ccccacctac 1440 cccttccttc ctctttatct ctcccacatt gtcttgctaa atatagactt ggtaattaaa 1500 atgttgattg aagtctggaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaa 1554 <210> 272 <211> 1511 <212> DNA <213> Homo sapiens <400> 272 ggggacccgc gggtttgcta tggcgatgag cagcggcggc agtggtggcg gcgtcccgga 60 gcaggaggat tccgtgctgt tccggcgcgg cacaggccag agcgatgatt ctgacatttg 120 ggatgataca gcactgataa aagcatatga taaagctgtg gcttcattta agcatgctct 180 aaagaatggt gacatttgtg aaacttcggg taaaccaaaa accacaccta aaagaaaacc 240 tgctaagaag aataaaagcc aaaagaagaa tactgcagct tccttacaac agtggaaagt 300 tggggacaaa tgttctgcca tttggtcaga agacggttgc atttacccag ctaccattgc 360 ttcaattgat tttaagagag aaacctgtgt tgtggtttac actggatatg gaaatagaga 420 ggagcaaaat ctgtccgatc tactttcccc aatctgtgaa gtagctaata atatagaaca 480 aaatgctcaa gagaatgaaa atgaaagcca agtttcaaca gatgaaagtg agaactccag 540 gtctcctgga aataaatcag ataacatcaa gcccaaatct gctccatgga actcttttct 600 ccctccacca ccccccatgc cagggccaag actgggacca ggaaagccag gtctaaaatt 660 caatggccca ccaccgccac cgccaccacc accaccccac ttactatcat gctggctgcc 720 tccatttcct tctggaccac caataattcc cccaccacct cccatatgtc cagattctct 780 tgatgatgct gatgctttgg gaagtatgtt aatttcatgg tacatgagtg gctatcatac 840 tggctattat atgggtttca gacaaaatca aaaagaagga aggtgctcac attccttaaa 900 ttaaggagaa atgctggcat agagcagcac taaatgacac cactaaagaa acgatcagac 960 agatctggaa tgtgaagcgt tatagaagat aactggcctc atttcttcaa aatatcaagt 1020 gttgggaaag aaaaaaggaa gtggaatggg taactcttct tgattaaaag ttatgtaata 1080 accaaatgca atgtgaaata ttttactgga ctctattttg aaaaaccatc tgtaaaagac 1140 tggggtgggg gtgggaggcc agcacggtgg tgaggcagtt gagaaaattt gaatgtggat 1200 tagattttga atgatattgg ataattattg gtaattttta tgagctgtga gaagggtgtt 1260 gtagtttata aaagactgtc ttaatttgca tacttaagca tttaggaatg aagtgttaga 1320 gtgtcttaaa atgtttcaaa tggtttaaca aaatgtatgt gaggcgtatg tggcaaaatg 1380 ttacagaatc taactggtgg acatggctgt tcattgtact gtttttttct atcttctata 1440 tgtttaaaag tatataataa aaatatttaa ttttttttta aaaaaaaaaa aaaaaaaaca 1500 aaaaaaaaaa a 1511 <210> 273 <211> 1300 <212> DNA <213> Homo sapiens <400> 273 ctgcagccgg tgcagttaca cgttttcctc caaggagcct cggacgttgt cacgggtttg 60 gggtcgggga cagagcggtg accatggcca ggctggcgtt gtctcctgtg cccagccact 120 ggatggtggc gttgctgctg ctgctctcag ctgagccagt accagcagcc agatcggagg 180 accggtaccg gaatcccaaa ggtagtgctt gttcgcggat ctggcagagc ccacgtttca 240 tagccaggaa acggggcttc acggtgaaaa tgcactgcta catgaacagc gcctccggca 300 atgtgagctg gctctggaag caggagatgg acgagaatcc ccagcagctg aagctggaaa 360 agggccgcat ggaagagtcc cagaacgaat ctctcgccac cctcaccatc caaggcatcc 420 ggtttgagga caatggcatc tacttctgtc agcagaagtg caacaacacc tcggaggtct 480 accagggctg cggcacagag ctgcgagtca tgggattcag caccttggca cagctgaagc 540 agaggaacac gctgaaggat ggtatcatca tgatccagac gctgctgatc atcctcttca 600 tcatcgtgcc tatcttcctg ctgctggaca aggatgacag caaggctggc atggaggaag 660 atcacaccta cgagggcctg gacattgacc agacagccac ctatgaggac atagtgacgc 720 tgcggacagg ggaagtgaag tggtctgtag gtgagcaccc aggccaggag tgagagccag 780 gtcgccccat gacctgggtg caggctccct ggcctcagtg actgcttcgg agctgcctgg 840 ctcatggccc aacccctttc ctggaccccc cagctggcct ctgaagctgg cccaccagag 900 ctgccatttg tctccagccc ctggtcccca gctcttgcca aagggcctgg agtagaagga 960 caacagggca gcaacttgga gggagttctc tggggatgga cgggacccag ccttctgggg 1020 gtgctatgag gtgatccgtc cccacacatg ggatggggga ggcagagact ggtccagagc 1080 ccgcaaatgg actcggagcc gagggcctcc cagcagagct tgggaagggc catggaccca 1140 actgggcccc agaagagcca caggaacatc attcctctcc cgcaaccact cccaccccag 1200 ggaggccctg gcctccagtg ccttcccccg tggaataaac ggtgtgtcct gagaaaccac 1260 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa 1300 <210> 274 <211> 5748 <212> DNA <213> Homo sapiens <400> 274 gagaagaaag ccagtgcgtc tctgggcgca ggggccagtg gggctcggag gcacaggcac 60 cccgcgacac tccaggttcc ccgacccacg tccctggcag ccccgattat ttacagcctc 120 agcagagcac ggggcggggg cagaggggcc cgcccgggag ggctgctact tcttaaaacc 180 tctgcgggct gcttagtcac agcccccctt gcttgggtgt gtccttcgct cgctccctcc 240 ctccgtctta ggtcactgtt ttcaacctcg aataaaaact gcagccaact tccgaggcag 300 cctcattgcc cagcggaccc cagcctctgc caggttcggt ccgccatcct cgtcccgtcc 360 tccgccggcc cctgccccgc gcccagggat cctccagctc ctttcgcccg cgccctccgt 420 tcgctccgga caccatggac aagttttggt ggcacgcagc ctggggactc tgcctcgtgc 480 cgctgagcct ggcgcagatc gatttgaata taacctgccg ctttgcaggt gtattccacg 540 tggagaaaaa tggtcgctac agcatctctc ggacggaggc cgctgacctc tgcaaggctt 600 tcaatagcac cttgcccaca atggcccaga tggagaaagc tctgagcatc ggatttgaga 660 cctgcaggta tgggttcata gaagggcacg tggtgattcc ccggatccac cccaactcca 720 tctgtgcagc aaacaacaca ggggtgtaca tcctcacatc caacacctcc cagtatgaca 780 catattgctt caatgcttca gctccacctg aagaagattg tacatcagtc acagacctgc 840 ccaatgcctt tgatggacca attaccataa ctattgttaa ccgtgatggc acccgctatg 900 tccagaaagg agaatacaga acgaatcctg aagacatcta ccccagcaac cctactgatg 960 atgacgtgag cagcggctcc tccagtgaaa ggagcagcac ttcaggaggt tacatctttt 1020 acaccttttc tactgtacac cccatcccag acgaagacag tccctggatc accgacagca 1080 cagacagaat ccctgctacc actttgatga gcactagtgc tacagcaact gagacagcaa 1140 ccaagaggca agaaacctgg gattggtttt catggttgtt tctaccatca gagtcaaaga 1200 atcatcttca cacaacaaca caaatggctg gtacgtcttc aaataccatc tcagcaggct 1260 gggagccaaa tgaagaaaat gaagatgaaa gagacagaca cctcagtttt tctggatcag 1320 gcattgatga tgatgaagat tttatctcca gcaccatttc aaccacacca cgggcttttg 1380 accacacaaa acagaaccag gactggaccc agtggaaccc aagccattca aatccggaag 1440 tgctacttca gacaaccaca aggatgactg atgtagacag aaatggcacc actgcttatg 1500 aaggaaactg gaacccagaa gcacaccctc ccctcattca ccatgagcat catgaggaag 1560 aagagacccc acattctaca agcacaatcc aggcaactcc tagtagtaca acggaagaaa 1620 cagctaccca gaaggaacag tggtttggca acagatggca tgagggatat cgccaaacac 1680 ccaaagaaga ctcccattcg acaacaggga cagctgcagc ctcagctcat accagccatc 1740 caatgcaagg aaggacaaca ccaagcccag aggacagttc ctggactgat ttcttcaacc 1800 caatctcaca ccccatggga cgaggtcatc aagcaggaag aaggatggat atggactcca 1860 gtcatagtat aacgcttcag cctactgcaa atccaaacac aggtttggtg gaagatttgg 1920 acaggacagg acctctttca atgacaacgc agcagagtaa ttctcagagc ttctctacat 1980 cacatgaagg cttggaagaa gataaagacc atccaacaac ttctactctg acatcaagca 2040 ataggaatga tgtcacaggt ggaagaagag acccaaatca ttctgaaggc tcaactactt 2100 tactggaagg ttatacctct cattacccac acacgaagga aagcaggacc ttcatcccag 2160 tgacctcagc taagactggg tcctttggag ttactgcagt tactgttgga gattccaact 2220 ctaatgtcaa tcgttcctta tcaggagacc aagacacatt ccaccccagt ggggggtccc 2280 ataccactca tggatctgaa tcagatggac actcacatgg gagtcaagaa ggtggagcaa 2340 acacaacctc tggtcctata aggacacccc aaattccaga atggctgatc atcttggcat 2400 ccctcttggc cttggctttg attcttgcag tttgcattgc agtcaacagt cgaagaaggt 2460 gtgggcagaa gaaaaagcta gtgatcaaca gtggcaatgg agctgtggag gacagaaagc 2520 caagtggact caacggagag gccagcaagt ctcaggaaat ggtgcatttg gtgaacaagg 2580 agtcgtcaga aactccagac cagtttatga cagctgatga gacaaggaac ctgcagaatg 2640 tggacatgaa gattggggtg taacacctac accattatct tggaaagaaa caaccgttgg 2700 aaacataacc attacaggga gctgggacac ttaacagatg caatgtgcta ctgattgttt 2760 cattgcgaat cttttttagc ataaaatttt ctactctttt tgttttttgt gttttgttct 2820 ttaaagtcag gtccaatttg taaaaacagc attgctttct gaaattaggg cccaattaat 2880 aatcagcaag aatttgatcg ttccagttcc cacttggagg cctttcatcc ctcgggtgtg 2940 ctatggatgg cttctaacaa aaactacaca tatgtattcc tgatcgccaa cctttccccc 3000 accagctaag gacatttccc agggttaata gggcctggtc cctgggagga aatttgaatg 3060 ggtccatttt gcccttccat agcctaatcc ctgggcattg ctttccactg aggttggggg 3120 ttggggtgta ctagttacac atcttcaaca gaccccctct agaaattttt cagatgcttc 3180 tgggagacac ccaaagggtg aagctattta tctgtagtaa actatttatc tgtgtttttg 3240 aaatattaaa ccctggatca gtcctttgat cagtataatt ttttaaagtt actttgtcag 3300 aggcacaaaa gggtttaaac tgattcataa taaatatctg tacttcttcg atcttcacct 3360 tttgtgctgt gattcttcag tttctaaacc agcactgtct gggtccctac aatgtatcag 3420 gaagagctga gaatggtaag gagactcttc taagtcttca tctcagagac cctgagttcc 3480 cactcagacc cactcagcca aatctcatgg aagaccaagg agggcagcac tgtttttgtt 3540 ttttgttttt tgtttttttt ttttgacact gtccaaaggt tttccatcct gtcctggaat 3600 cagagttgga agctgaggag cttcagcctc ttttatggtt taatggccac ctgttctctc 3660 ctgtgaaagg ctttgcaaag tcacattaag tttgcatgac ctgttatccc tggggcccta 3720 tttcatagag gctggcccta ttagtgattt ccaaaaacaa tatggaagtg ccttttgatg 3780 tcttacaata agagaagaag ccaatggaaa tgaaagagat tggcaaaggg gaaggatgat 3840 gccatgtaga tcctgtttga catttttatg gctgtatttg taaacttaaa cacaccagtg 3900 tctgttcttg atgcagttgc tatttaggat gagttaagtg cctggggagt ccctcaaaag 3960 gttaaaggga ttcccatcat tggaatctta tcaccagata ggcaagttta tgaccaaaca 4020 agagagtact ggctttatcc tctaacctca tattttctcc cacttggcaa gtcctttgtg 4080 gcatttattc atcagtcagg gtgtccgatt ggtcctagaa cttccaaagg ctgcttgtca 4140 tagaagccat tgcatctata aagcaacggc tcctgttaaa tggtatctcc tttctgaggc 4200 tcctactaaa agtcatttgt tacctaaact tatgtgctta acaggcaatg cttctcagac 4260 cacaaagcag aaagaagaag aaaagctcct gactaaatca gggctgggct tagacagagt 4320 tgatctgtag aatatcttta aaggagagat gtcaactttc tgcactattc ccagcctctg 4380 ctcctccctg tctaccctct cccctccctc tctccctcca cttcacccca caatcttgaa 4440 aaacttcctt tctcttctgt gaacatcatt ggccagatcc attttcagtg gtctggattt 4500 ctttttattt tcttttcaac ttgaaagaaa ctggacatta ggccactatg tgttgttact 4560 gccactagtg ttcaagtgcc tcttgttttc ccagagattt cctgggtctg ccagaggccc 4620 agacaggctc actcaagctc tttaactgaa aagcaacaag ccactccagg acaaggttca 4680 aaatggttac aacagcctct acctgtcgcc ccagggagaa aggggtagtg atacaagtct 4740 catagccaga gatggttttc cactccttct agatattccc aaaaagaggc tgagacagga 4800 ggttattttc aattttattt tggaattaaa tacttttttc cctttattac tgttgtagtc 4860 cctcacttgg atatacctct gttttcacga tagaaataag ggaggtctag agcttctatt 4920 ccttggccat tgtcaacgga gagctggcca agtcttcaca aacccttgca acattgcctg 4980 aagtttatgg aataagatgt attctcactc ccttgatctc aagggcgtaa ctctggaagc 5040 acagcttgac tacacgtcat ttttaccaat gattttcagg tgacctgggc taagtcattt 5100 aaactgggtc tttataaaag taaaaggcca acatttaatt attttgcaaa gcaacctaag 5160 agctaaagat gtaatttttc ttgcaattgt aaatcttttg tgtctcctga agacttccct 5220 taaaattagc tctgagtgaa aaatcaaaag agacaaaaga catcttcgaa tccatatttc 5280 aagcctggta gaattggctt ttctagcaga acctttccaa aagttttata ttgagattca 5340 taacaacacc aagaattgat tttgtagcca acattcattc aatactgtta tatcagagga 5400 gtaggagaga ggaaacattt gacttatctg gaaaagcaaa atgtacttaa gaataagaat 5460 aacatggtcc attcaccttt atgttataga tatgtctttg tgtaaatcat ttgttttgag 5520 ttttcaaaga atagcccatt gttcattctt gtgctgtaca atgaccactg ttattgttac 5580 tttgactttt cagagcacac ccttcctctg gtttttgtat atttattgat ggatcaataa 5640 taatgaggaa agcatgatat gtatattgct gagttgaaag cacttattgg aaaatattaa 5700 aaggctaaca ttaaaagact aaaggaaaca gaaaaaaaaa aaaaaaaa 5748 <210> 275 <211> 1924 <212> DNA <213> Homo sapiens <400> 275 cgtagctatt tcaaggcgcg cgcctcgtgg tggactcacc gctagcccgc agcgctcggc 60 ttcctggtaa ttcttcacct cttttctcag ctccctgcag catgggtgct gggccctcct 120 tgctgctcgc cgccctcctg ctgcttctct ccggcgacgg cgccgtgcgc tgcgacacac 180 ctgccaactg cacctatctt gacctgctgg gcacctgggt cttccaggtg ggctccagcg 240 gttcccagcg cgatgtcaac tgctcggtta tgggaccaca agaaaaaaaa gtagtggtgt 300 accttcagaa gctggataca gcatatgatg accttggcaa ttctggccat ttcaccatca 360 tttacaacca aggctttgag attgtgttga atgactacaa gtggtttgcc ttttttaagt 420 ataaagaaga gggcagcaag gtgaccactt actgcaacga gacaatgact gggtgggtgc 480 atgatgtgtt gggccggaac tgggcttgtt tcaccggaaa gaaggtggga actgcctctg 540 agaatgtgta tgtcaacata gcacacctta agaattctca ggaaaagtat tctaataggc 600 tctacaagta tgatcacaac tttgtgaaag ctatcaatgc cattcagaag tcttggactg 660 caactacata catggaatat gagactctta ccctgggaga tatgattagg agaagtggtg 720 gccacagtcg aaaaatccca aggcccaaac ctgcaccact gactgctgaa atacagcaaa 780 agattttgca tttgccaaca tcttgggact ggagaaatgt tcatggtatc aattttgtca 840 gtcctgttcg aaaccaagca tcctgtggca gctgctactc atttgcttct atgggtatgc 900 tagaagcgag aatccgtata ctaaccaaca attctcagac cccaatccta agccctcagg 960 aggttgtgtc ttgtagccag tatgctcaag gctgtgaagg cggcttccca taccttattg 1020 caggaaagta cgcccaagat tttgggctgg tggaagaagc ttgcttcccc tacacaggca 1080 ctgattctcc atgcaaaatg aaggaagact gctttcgtta ttactcctct gagtaccact 1140 atgtaggagg tttctatgga ggctgcaatg aagccctgat gaagcttgag ttggtccatc 1200 atgggcccat ggcagttgct tttgaagtat atgatgactt cctccactac aaaaagggga 1260 tctaccacca cactggtcta agagaccctt tcaacccctt tgagctgact aatcatgctg 1320 ttctgcttgt gggctatggc actgactcag cctctgggat ggattactgg attgttaaaa 1380 acagctgggg caccggctgg ggtgagaatg gctacttccg gatccgcaga ggaactgatg 1440 agtgtgcaat tgagagcata gcagtggcag ccacaccaat tcctaaattg tagggtatgc 1500 cttccagtat ttcataatga tctgcatcag ttgtaaaggg gaattggtat attcacagac 1560 tgtagacttt cagcagcaat ctcagaagct tacaaataga tttccatgaa gatatttgtc 1620 ttcagaatta aaactgccct taattttaat atacctttca atcggccact ggccattttt 1680 ttctaagtat tcaattaagt gggaattttc tggaagatgg tcagctatga agtaatagag 1740 tttgcttaat catttgtaat tcaaacatgc tatatttttt aaaatcaatg tgaaaacata 1800 gacttatttt taaattgtac caatcacaag aaaataatgg caataattat caaaactttt 1860 aaaatagatg ctcatatttt taaaataaag ttttaaaaat aactgcaaaa aaaaaaaaaa 1920 aaaa 1924 <210> 276 <211> 2344 <212> DNA <213> Homo sapiens <400> 276 cggccgcctc cgcgtccgcg tcgtcgtctg tgctcccggc gctgacgtgt ctgggcggtc 60 ggcttccact ccttcaggcg tcggcagcca ctagtcgtgg cgagaggggc ggggtggccg 120 gggctggcgc tccacttggc ccccgctccc ggcccgcccc gccgccgcgg ccccccggat 180 gagggtatat attcggagcg agcgcgggac gccgatgagt ggccgcgcgg aaggagctgg 240 agacggtcgt agctgcggtc gcgccgagaa aggtttacag gtacatacat tacaccccta 300 tttctacaaa gcttggctat tagagcatta tgaacattaa tgacctcaaa ctcacgttgt 360 ccaaagctgg gcaagagcac ctactacgtt tctggaatga gcttgaagaa gcccaacagg 420 tagaacttta tgcagagctc caggccatga actttgagga gctgaacttc tttttccaaa 480 aggccattga aggttttaac cagtcttctc accaaaagaa tgtggatgca cgaatggaac 540 ctgtgcctcg agaggtatta ggcagtgcta caagggatca agatcagctc caggcctggg 600 aaagtgaagg acttttccag atttctcaga ataaagtagc agttcttctt ctagctggtg 660 ggcaggggac aagactcggc gttgcatatc ctaaggggat gtatgatgtt ggtttgccat 720 cccgtaagac actttttcag attcaagcag agcgtatcct gaagctacag caggttgctg 780 aaaaatatta tggcaacaaa tgcattattc catggtatat aatgaccagt ggcagaacaa 840 tggaatctac aaaggagttc ttcaccaagc acaagtactt tggtttaaaa aaagagaatg 900 taatcttttt tcagcaagga atgctccccg ccatgagttt tgatgggaaa attattttgg 960 aagagaagaa caaagtttct atggctccag atgggaatgg tggtctttat cgggcacttg 1020 cagcccagaa tattgtggag gatatggagc aaagaggcat ttggagcatt catgtctatt 1080 gtgttgacaa catattagta aaagtggcag acccacggtt cattggattt tgcattcaga 1140 aaggagcaga ctgtggagca aaggtggtag agaaaacgaa ccctacagaa ccagttggag 1200 tggtttgccg agtggatgga gtttaccagg tggtagaata tagtgagatt tccctggcaa 1260 cagctcaaaa acgaagctca gacggacgac tgctgttcaa tgcggggaac attgccaacc 1320 atttcttcac tgtaccattt ctgagagatg ttgtcaatgt ttatgaacct cagttgcagc 1380 accatgtggc tcaaaagaag attccttatg tggataccca aggacagtta attaagccag 1440 acaaacccaa tggaataaag atggaaaaat ttgtctttga catcttccag tttgcaaaga 1500 agtttgtggt atatgaagta ttgcgagaag atgagttttc cccactaaag aatgctgata 1560 gtcagaatgg gaaagacaac cctactactg caaggcatgc tttgatgtcc cttcatcatt 1620 gctgggtcct caatgcaggg ggccatttca tagatgaaaa tggctctcgc cttccagcaa 1680 ttccccgctt gaaggatgcc aatgatgtac caatccaatg tgaaatctct cctcttatct 1740 cctatgctgg agaaggatta gaaagttatg tggcagataa agaattccat gcacctctaa 1800 tcatcgatga gaatggagtt catgagctgg tgaaaaatgg tatttgaacc agataccaag 1860 ttttgtttgc cacgatagga atagctttta tttttgatag accaactgtg aacctacaag 1920 acgtcttgga caactgaagt ttaaatatcc acagggtttt attttgcttg ttgaactctt 1980 agagctattg caaacttccc aagatccaga tgactgaatt tcagatagca tttttatgat 2040 tcccaactca ttgaaggtct tatttatata attttttcca agccaaggag accattggcc 2100 atccaggaaa tttcgtacag ctgaaatata ggcaggatgt tcaacatcag tttacttgca 2160 gctggaagca tttgtttttg aagttgtaca tagtaataat atgtcattgt acatgttgaa 2220 aggtttctat ggtactaaaa gtttgtttta ttttatcaaa cattaagctt ttttaagaaa 2280 ataattgggc agtgaaataa atgtatcttc ttgtctctgg agtgtcaaaa aaaaaaaaaa 2340 aaaa 2344 <210> 277 <211> 3484 <212> DNA <213> Homo sapiens <400> 277 gtgcgagccc ggccgccggt gagtcggctg gagcgcatct ggtcctccgc gcggaaagcg 60 ctgcttttgc ctggccgccc tagccgctgg ctcatccaag tggccttcgc cgctctcttg 120 cgtcccaacc agagcgctgg ccacctcgcc gcccagctca cgccgcgccc gcgctcccag 180 gctccgggtt ttcttaaatg ttttcttgga gccttaaaga tggagatgac agaaatgact 240 ggtgtgtcgc tgaaacgtgg ggcactggtt gtcgaagata atgacagtgg agtcccagtt 300 gaagagacaa aaaaacagaa gctgtcggaa tgcagtctaa ccaaaggtca agatgggcta 360 cagaatgact ttctgtccat cagtgaagac gtgcctcggc ctcctgacac tgtcagtact 420 gggaaaggtg gaaagaattc tgaggctcag ttggaagatg aggaagaaga ggaggaagat 480 ggactttcag aggagtgcga ggaggaggaa tcagagagtt ttgcagacat gatgaagcat 540 ggactcactg aggctgacgt aggcatcacc aagtttgtga gttctcatca agggttctcg 600 ggaatcttaa aagaaagata ctccgacttc gttgttcatg aaataggaaa agatggacgg 660 atcagccatt tgaatgactt gtccattcca gtggatgagg aggacccttc agaagacata 720 tttacagttt tgacagctga agaaaagcag cgattggaag agctccagct gttcaaaaat 780 aaggaaacca gtgttgccat tgaggttatc gaggacacca aagagaaaag aaccatcatc 840 catcaggcta tcaaatctct gtttccagga ttagagacaa aaacagagga tagggagggg 900 aagaaataca ttgtagccta ccacgcagct gggaaaaagg ctttggcaaa tccaagaaaa 960 cattcttggc caaaatctag gggaagttac tgccacttcg tactatataa ggaaaacaaa 1020 gacaccatgg atgctattaa tgtactctcc aaatacttaa gagtcaagcc aaatatattc 1080 tcctacatgg gaaccaaaga taaaagggct ataacagttc aagaaattgc tgttctcaaa 1140 ataactgcac aaagacttgc ccacctgaat aagtgcttga tgaactttaa gctagggaat 1200 ttcagctatc aaaaaaaccc actgaaattg ggagagcttc aaggaaacca cttcactgtt 1260 gttctcagaa atataacagg aactgatgac caagtacagc aagctatgaa ctctctcaag 1320 gagattggat ttattaacta ctatggaatg caaagatttg gaaccacagc tgtccctacg 1380 tatcaggttg gaagagctat actacaaaat tcctggacag aagtcatgga tttaatattg 1440 aaaccccgct ctggagctga aaagggctac ttggttaaat gcagagaaga atgggcaaag 1500 accaaagacc caactgctgc cctcagaaaa ctacctgtca aaaggtgtgt ggaagggcag 1560 ctgcttcgag gactttcaaa atatggaatg aagaatatag tctctgcatt tggcataata 1620 cccagaaata atcgcttaat gtatattcat agctaccaaa gctatgtgtg gaataacatg 1680 gtaagcaaga ggatagaaga ctatggacta aaacctgttc caggggacct cgttctcaaa 1740 ggagccacag ccacctatat tgaggaagat gatgttaata attactctat ccatgatgtg 1800 gtaatgccct tgcctggttt cgatgttatc tacccaaagc ataaaattca agaagcctac 1860 agggaaatgc tcacagctga caatcttgat attgacaaca tgagacacaa aattcgagat 1920 tattccttgt caggggccta ccgaaagatc attattcgtc ctcagaatgt tagctgggaa 1980 gtcgttgcat atgatgatcc caaaattcca cttttcaaca cagatgtgga caacctagaa 2040 gggaagacac caccagtttt tgcttctgaa ggcaaataca gggctctgaa aatggatttt 2100 tctctacccc cttctactta cgccaccatg gccattcgag aagtgctaaa aatggatacc 2160 agtatcaaga accagacgca gctgaataca acctggcttc gctgagcagt accttgtcca 2220 cagattagaa aacgtacaca agtgtttgct tcctggctcc ctgtgcattt ttgtcttagt 2280 tcagactcat atatggattt caaatctttg taataaaaat tatttgtatt tttaagtttt 2340 tattagctta aagaaataat ttgcaatatt tgtacatgta cacaaatcct gaggttctta 2400 attttagctc agaatataaa ttagtcaaaa tacacttcag gtgcttaaat cagagtaaaa 2460 tgtcagcttt acaataataa aaaaaggact ttggtttaaa gtagcaggtt taggttttgc 2520 tacattctca aaagacagca ggagtatttg acacatctgt gatggagtat acaacaatgc 2580 attttaagag caaatgcaac aaaacaaatc tggactatgg ataaataatt tgagagctgc 2640 cacccacaaa tataaataca gtactcatgc tgactgaaat aataagacat ctacaaattt 2700 ataaacaaaa agtgattgtc attatcctgc ttatgtacta gattcaggca agcattatag 2760 actttttggt tgcggtggct tttgcattta tattatcaat gccttgcagg aacgttgcat 2820 tgataggccc attttatttt tttatttttt ttttcgagac aggatctcac tctgtagcac 2880 aggctggatt gcagtgcaat cctgcaattc tcaatcttgc actgcagcct cgacctccca 2940 ggctccagtg actctcccac ctcagcctcc taagtagctg ggagtacagg cgcgcaccac 3000 cacgcctagc tgatttttgt atttttttgt agagacgggg gtttggccat gttgccgagg 3060 ctaactcctg ggattacagg catgagctgt gctggccggg tttttttttc ttgatgtaaa 3120 cgtgtacagc tgttttatta gttaaggtct aatttttact ctaggtgcct tttatgttca 3180 gaactctttc cactggactg gtatttgctc aaaaataaat aatggtagag aagaaaacta 3240 taaaaatgga caaggctttc ttctatcagt agcgtttacc ctttgtcacc agtggctttg 3300 gtatttccat gtctggcatt gcataaactt ctctggtgtg aaaggataaa tatgcctttc 3360 taaagttgta tatcaaaatt gtatcaattt ttattttcta tgatttctag aaacaaatgt 3420 aataaatatt tttaaaatct cctttctact ggttatgtaa ataaatcaaa taaatatatc 3480 aaaa 3484 <210> 278 <211> 1498 <212> DNA <213> Homo sapiens <400> 278 gaggccagag tgccatcgaa ggtaattata gagacagtaa aatcctttta ctctgggaaa 60 aataaaatgc tgggtgtctc acaaaatttc agaacctgat ttcaaacgga tcataacaaa 120 gaggagatca aatttagcat ggtggactgc tcgacaggat atatttgtca atggaatgtt 180 tccacatatt ataccaccaa catgagaaaa aaatgatcat tgtttatttg aagcttgatg 240 atattctaac gctgcctttt ctcttctcat tttagagaaa aatgagcagg cggaattgtt 300 ggatttgtaa gatgtgcaga gatgaatcta agaggccccc ttcaaacctt actttggagg 360 aagtattaca gtgggcccag tcttttgaaa atttaatggc tacaaaatat ggtccagtag 420 tctatgcagc atatttaaaa atggagcaca gtgacgagaa tattcaattc tggatggcat 480 gtgaaaccta taagaaaatt gcctcacggt ggagcagaat ttctagggca aagaagcttt 540 ataagattta catccagcca cagtccccta gagagattaa cattgacagt tcgacaagag 600 agactatcat caggaacatt caggaaccca ctgaaacatg ttttgaagaa gctcagaaaa 660 tagtctatat gcatatggaa agggattcct accccagatt tctaaagtca gaaatgtacc 720 aaaaactttt gaaaactatg cagtccaaca acagtttctg actacaactc aaaagtttaa 780 atagaaaaca gtatattgaa agtggtgggt ttgatctttt tatttagaaa cccacaaaat 840 cagaaacaca gtacaaataa aacagaaatc aaactataag ttgactttta gttcctaaaa 900 agaaacatat ttcaaaagca atggaatcta gaattcttat aacatgaata acaaaatgta 960 cagcaagcct atgtagttca attaatatat aaggaaaagg aaggtctttc ttcatgatac 1020 aagcattata aagtttttac tgtagtagtc aattaatgga tatttccttg ttaataaaat 1080 tttgtgtcat aatttacaaa ttagttcttt aaaaattgtt gttatatgaa ttgtgtttct 1140 agcatgaatg ttctatagag tactctaaat aacttgaatt tatagacaaa tgctactcac 1200 agtacaatca attgtattat accatgagaa aatcaaaaag gtgttcttca gagacatttt 1260 atctataaaa ttttcctact attatgttca ttaacaaact tctttatcac atgtatcttc 1320 tacatgtaaa acatttctga tgatttttta acaaaaaata tatgaatttc ttcatttgct 1380 cttgcatcta cattgctata aggatataaa atgtggtttc tatattttga gatgtttttt 1440 ccttacaatg tgaactcatc gtgatcttgg aaatcaataa agtcaaatat caactaaa 1498 <210> 279 <211> 3293 <212> DNA <213> Homo sapiens <400> 279 cttttgctct cagatgctgc cagggtccct gaagagggaa gacacgcgga aacaggcttg 60 cacccagaca cgacaccatg catctcctcg gcccctggct cctgctcctg gttctagaat 120 acttggcttt ctctgactca agtaaatggg tttttgagca ccctgaaacc ctctacgcct 180 gggagggggc ctgcgtctgg atcccctgca cctacagagc cctagatggt gacctggaaa 240 gcttcatcct gttccacaat cctgagtata acaagaacac ctcgaagttt gatgggacaa 300 gactctatga aagcacaaag gatgggaagg ttccttctga gcagaaaagg gtgcaattcc 360 tgggagacaa gaataagaac tgcacactga gtatccaccc ggtgcacctc aatgacagtg 420 gtcagctggg gctgaggatg gagtccaaga ctgagaaatg gatggaacga atacacctca 480 atgtctctga aaggcctttt ccacctcata tccagctccc tccagaaatt caagagtccc 540 aggaagtcac tctgacctgc ttgctgaatt tctcctgcta tgggtatccg atccaattgc 600 agtggctcct agagggggtt ccaatgaggc aggctgctgt cacctcgacc tccttgacca 660 tcaagtctgt cttcacccgg agcgagctca agttctcccc acagtggagt caccatggga 720 agattgtgac ctgccagctt caggatgcag atgggaagtt cctctccaat gacacggtgc 780 agctgaacgt gaagcacacc ccgaagttgg agatcaaggt cactcccagt gatgccatag 840 tgagggaggg ggactctgtg accatgacct gcgaggtcag cagcagcaac ccggagtaca 900 cgacggtatc ctggctcaag gatgggacct cgctgaagaa gcagaataca ttcacgctaa 960 acctgcgcga agtgaccaag gaccagagtg ggaagtactg ctgtcaggtc tccaatgacg 1020 tgggcccggg aaggtcggaa gaagtgttcc tgcaagtgca gtatgccccg gaaccttcca 1080 cggttcagat cctccactca ccggctgtgg agggaagtca agtcgagttt ctttgcatgt 1140 cactggccaa tcctcttcca acaaattaca cgtggtacca caatgggaaa gaaatgcagg 1200 gaaggacaga ggagaaagtc cacatcccaa agatcctccc ctggcacgct gggacttatt 1260 cctgtgtggc agaaaacatt cttggtactg gacagagggg cccgggagct gagctggatg 1320 tccagtatcc tcccaagaag gtgaccacag tgattcaaaa ccccatgccg attcgagaag 1380 gagacacagt gaccctttcc tgtaactaca attccagtaa ccccagtgtt acccggtatg 1440 aatggaaacc ccatggcgcc tgggaggagc catcgcttgg ggtgctgaag atccaaaacg 1500 ttggctggga caacacaacc atcgcctgcg cagcttgtaa tagttggtgc tcgtgggcct 1560 cccctgtcgc cctgaatgtc cagtatgccc cccgagacgt gagggtccgg aaaatcaagc 1620 ccctttccga gattcactct ggaaactcgg tcagcctcca atgtgacttc tcaagcagcc 1680 accccaaaga agtccagttc ttctgggaga aaaatggcag gcttctgggg aaagaaagcc 1740 agctgaattt tgactccatc tccccagaag atgctgggag ttacagctgc tgggtgaaca 1800 actccatagg acagacagcg tccaaggcct ggacacttga agtgctgtat gcacccagga 1860 ggctgcgtgt gtccatgagc ccgggggacc aagtgatgga ggggaagagt gcaaccctga 1920 cctgtgagag cgacgccaac cctcccgtct cccactacac ctggtttgac tggaataacc 1980 aaagcctccc ctaccacagc cagaagctga gattggagcc ggtgaaggtc cagcactcgg 2040 gtgcctactg gtgccagggg accaacagtg tgggcaaggg ccgttcgcct ctcagcaccc 2100 tcaccgtcta ctatagcccg gagaccatcg gcaggcgagt ggctgtggga ctcgggtcct 2160 gcctcgccat cctcatcctg gcaatctgtg ggctcaagct ccagcgacgt tggaagagga 2220 cacagagcca gcaggggctt caggagaatt ccagcggcca gagcttcttt gtgaggaata 2280 aaaaggttag aagggccccc ctctctgaag gcccccactc cctgggatgc tacaatccaa 2340 tgatggaaga tggcattagc tacaccaccc tgcgctttcc cgagatgaac ataccacgaa 2400 ctggagatgc agagtcctca gagatgcaga gacctccccc ggactgcgat gacacggtca 2460 cttattcagc attgcacaag cgccaagtgg gcgactatga gaacgtcatt ccagattttc 2520 cagaagatga ggggattcat tactcagagc tgatccagtt tggggtcggg gagcggcctc 2580 aggcacaaga aaatgtggac tatgtgatcc tcaaacattg acactggatg ggctgcagca 2640 gaggcactgg gggcagcggg ggccagggaa gtccccgagt ttccccagac accgccacat 2700 ggcttcctcc tgcgcgcatg tgcgcacaca cacacacaca cgcacacaca cacacacaca 2760 ctcactgcgg agaaccttgt gcctggctca gagccagtct ttttggtgag ggtaacccca 2820 aacctccaaa actcctgccc ctgttctctt ccactctcct tgctacccag aaatccatct 2880 aaatacctgc cctgacatgc acacctcccc ctgcccccac cacggccact ggccatctcc 2940 acccccagct gcttgtgtcc ctcctgggat ctgctcgtca tcatttttcc ttcccttctc 3000 catctctctg gccctctacc cctgatctga catccccact cacgaatatt atgcccagtt 3060 tctgcctctg agggaaagcc cagaaaagga cagaaacgaa gtagaaaggg gcccagtcct 3120 ggcctggctt ctcctttgga agtgaggcat tgcacgggga gacgtacgta tcagcggccc 3180 cttgactctg gggactccgg gtttgagatg gacacactgg tgtggattaa cctgccaggg 3240 agacagagct cacaataaaa atggctcaga tgccacttca aagaaaaaaa aaa 3293 <210> 280 <211> 1621 <212> DNA <213> Homo sapiens <400> 280 ccacaaatgt gggagggcga taaccactcg tagaaagcgt gagaagttac tacaagcggt 60 cctcccggcc accgtactgt tccgctccca gaagccccgg gcggcggaag tcgtcactct 120 taagaaggga cggggcccca cgctgcgcac ccgcgggttt gctatggcga tgagcagcgg 180 cggcagtggt ggcggcgtcc cggagcagga ggattccgtg ctgttccggc gcggcacagg 240 ccagagcgat gattctgaca tttgggatga tacagcactg ataaaagcat atgataaagc 300 tgtggcttca tttaagcatg ctctaaagaa tggtgacatt tgtgaaactt cgggtaaacc 360 aaaaaccaca cctaaaagaa aacctgctaa gaagaataaa agccaaaaga agaatactgc 420 agcttcctta caacagtgga aagttgggga caaatgttct gccatttggt cagaagacgg 480 ttgcatttac ccagctacca ttgcttcaat tgattttaag agagaaacct gtgttgtggt 540 ttacactgga tatggaaata gagaggagca aaatctgtcc gatctacttt ccccaatctg 600 tgaagtagct aataatatag aacagaatgc tcaagagaat gaaaatgaaa gccaagtttc 660 aacagatgaa agtgagaact ccaggtctcc tggaaataaa tcagataaca tcaagcccaa 720 atctgctcca tggaactctt ttctccctcc accacccccc atgccagggc caagactggg 780 accaggaaag ccaggtctaa aattcaatgg cccaccaccg ccaccgccac caccaccacc 840 ccacttacta tcatgctggc tgcctccatt tccttctgga ccaccaataa ttcccccacc 900 acctcccata tgtccagatt ctcttgatga tgctgatgct ttgggaagta tgttaatttc 960 atggtacatg agtggctatc atactggcta ttatatgggt ttcagacaaa atcaaaaaga 1020 aggaaggtgc tcacattcct taaattaagg agaaatgctg gcatagagca gcactaaatg 1080 acaccactaa agaaacgatc agacagatct ggaatgtgaa gcgttataga agataactgg 1140 cctcatttct tcaaaatatc aagtgttggg aaagaaaaaa ggaagtggaa tgggtaactc 1200 ttcttgatta aaagttatgt aataaccaaa tgcaatgtga aatattttac tggactcttt 1260 tgaaaaacca tctgtaaaag actggggtgg gggtgggagg ccagcacggt ggtgaggcag 1320 ttgagaaaat ttgaatgtgg attagatttt gaatgatatt ggataattat tggtaatttt 1380 atggcctgtg agaagggtgt tgtagtttat aaaagactgt cttaatttgc atacttaagc 1440 atttaggaat gaagtgttag agtgtcttaa aatgtttcaa atggtttaac aaaatgtatg 1500 tgaggcgtat gtggcaaaat gttacagaat ctaactggtg gacatggctg ttcattgtac 1560 tgtttttttc tatcttctat atgtttaaaa gtatataata aaaatattta attttttttt 1620 a 1621 <210> 281 <211> 1572 <212> DNA <213> Homo sapiens <400> 281 aagttgcttt tgtccaaaca tccgggcttc tcctttttgt gttccggccg atcccacctc 60 tcctcgaccc tggacgtcta ccttccggag gcccacatct tgcccactcc gcgcgcgggg 120 ctagcgcggg tttcagcgac gggagccctc aagggacatg gcaactacag cggcgccggc 180 gggcggcgcc cgaaatggag ctggcccgga atggggaggg ttcgaagaaa acatccaggg 240 cggaggctca gctgtgattg acatggagaa catggatgat acctcaggct ctagcttcga 300 ggatatgggt gagctgcatc agcgcctgcg cgaggaagaa gtagacgctg atgcagctga 360 tgcagctgct gctgaagagg aggatggaga gttcctgggc atgaagggct ttaagggaca 420 gctgagccgg caggtggcag atcagatgtg gcaggctggg aaaagacaag cctccagggc 480 cttcagcttg tacgccaaca tcgacatcct cagaccctac tttgatgtgg agcctgctca 540 ggtgcgaagc aggctcctgg agtccatgat ccctatcaag atggtcaact tcccccagaa 600 aattgcaggt gaactctatg gacctctcat gctggtcttc actctggttg ctatcctact 660 ccatgggatg aagacgtctg acactattat ccgggagggc accctgatgg gcacagccat 720 tggcacctgc ttcggctact ggctgggagt ctcatccttc atttacttcc ttgcctacct 780 gtgcaacgcc cagatcacca tgctgcagat gttggcactg ctgggctatg gcctctttgg 840 gcattgcatt gtcctgttca tcacctataa tatccacctc cacgccctct tctacctctt 900 ctggctgttg gtgggtggac tgtccacact gcgcatggta gcagtgttgg tgtctcggac 960 cgtgggcccc acacagcggc tgctcctctg tggcaccctg gctgccctac acatgctctt 1020 cctgctctat ctgcattttg cctaccacaa agtggtagag gggatcctgg acacactgga 1080 gggccccaac atcccgccca tccagagggt ccccagagac atccctgcca tgctccctgc 1140 tgctcggctt cccaccaccg tcctcaacgc cacagccaaa gctgttgcgg tgaccctgca 1200 gtcacactga ccccacctga aattcttggc cagtcctctt tcccgcagct gcagagagga 1260 ggaagactat taaaggacag tcctgatgac atgtttcgta gatggggttt gcagctgcca 1320 ctgagctgta gctgcgtaag tacctccttg atgcctgtcg gcacttctga aaggcacaag 1380 gccaagaact cctggccagg actgcaaggc tctgcagcca atgcagaaaa tgggtcagct 1440 cctttgagaa cccctcccca cctacccctt ccttcctctt tatctctccc acattgtctt 1500 gctaaatata gacttggtaa ttaaaatgtt gattgaagtc tggaactgca aaaaaaaaaa 1560 aaaccaaaaa aa 1572

Claims (55)

(a) measuring the expression level of one or more marker genes in a sample comprising B lymphoma cells obtained from a subject, wherein the one or more marker genes are selected from IFITM1, CD40, RGS13, VNN2, LMO2, CD79B, CD22, BTG2, IGF1R, CD44, CTSC, EPDR1, UAP1, and PUS7;
(b) predicting whether the subject will respond to anti-CD40 antibody treatment based on the measured expression levels of the one or more marker genes from step (a)
A method for predicting responsiveness of a subject with B-cell lymphoma to anti-CD40 antibody treatment comprising a. The method of claim 1, wherein the measured expression level is normalized. The method of claim 1 or 2, wherein the anti-CD40 antibody treatment is treatment with an agonist anti-CD40 antibody, and increased expression of one or more of IFITM1, CD79B, IGF1R, CD44, CTSC, EPDR1, and PUS7 relative to the reference level. Is indicative of whether said subject will respond less to agonist anti-CD40 antibody treatment. The method of claim 3, wherein the reference level is determined based on the expression level of the corresponding marker gene in a sample comprising B lymphoma cells from a subject having increased tumor volume after anti-CD40 antibody treatment. The method of claim 4, wherein the sample from the subject for determining the reference level comprises B lymphoma cells of the same type as the sample from the subject for which responsiveness to anti-CD40 antibody treatment is predicted. The method of claim 1 or 2, wherein the anti-CD40 antibody treatment is treatment with an agonist anti-CD40 antibody and increased expression of one or more of CD40, RGS13, VNN2, LMO2, CD22, BTG2, and UAP1 relative to the reference level. Is indicative of whether the subject will respond to agonist anti-CD40 antibody treatment. The method of claim 6, wherein the reference level is determined based on the expression level of the corresponding marker gene in the sample comprising B lymphoma cells from a subject whose tumor volume is reduced after anti-CD40 antibody treatment. The method of claim 7, wherein the sample from the subject for determining the reference level comprises B lymphoma cells of the same type as the sample from the subject for which responsiveness to anti-CD40 antibody treatment is predicted. 9. The method of claim 3, wherein the agonist anti-CD40 antibody stimulates CD40 and enhances interaction between CD40 and CD40 ligand. 10. The method of claim 9, wherein the agonist anti-CD40 antibody comprises the heavy chain amino acid sequence set forth in SEQ ID NO: 1 and the light chain amino acid sequence set forth in SEQ ID NO: 2. 9. The method of claim 3, wherein the agonist anti-CD40 antibody stimulates CD40 and does not enhance or inhibit the interaction between CD40 and CD40 ligand. 10. The method of claim 1, wherein at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, or The expression level of 14 marker genes is measured. The method of claim 12, wherein the expression levels of IFITM1, RGS13, CD79B, CD22, BTG2, CD44, EPDR1, and UAP1 are measured. The method of claim 1, wherein the B cell lymphoma is diffuse large cell B-cell lymphoma (DLBCL). The method of any one of claims 1-13, wherein the B cell lymphoma is a non-Hodgkin's lymphoma. The method of claim 15, wherein the non-Hodgkin's lymphoma is follicular lymphoma, mantle cell lymphoma, marginal lymphoma, or small lymphocytic lymphoma. The method of claim 1, wherein the sample comprising B lymphoma cells is a formalin fixed paraffin embedded biopsy sample. The method of claim 1, wherein the expression level of one or more marker genes is measured by the level of RNA transcript of one or more marker genes. The method of claim 18, wherein the RNA transcript is measured by qRT-PCR. The method of claim 1, wherein the expression level of one or more marker genes is measured by the level of protein expression of one or more marker genes. The method of claim 1, further comprising measuring the expression level of BCL6, wherein a higher expression level of BCL6 relative to the reference level indicates whether the subject will respond to anti-CD40 antibody treatment. Method. The method of claim 21, wherein the reference level is determined based on the expression level of BCL6 in a sample comprising B lymphoma cells from a subject having reduced tumor volume after anti-CD40 antibody treatment. (a) selected from the group consisting of IFITM1, CD40, RGS13, VNN2, LMO2, CD79B, CD22, BTG2, IGF1R, CD44, CTSC, EPDR1, UAP1, PUS7, and BCL6 in a sample comprising B lymphoma cells obtained from a subject Determining the expression level of one or more marker genes;
(b) producing a report summarizing the expression levels of one or more marker genes obtained in step (a)
A method of creating a personalized genomics profile for a subject with B-cell lymphoma, comprising. The method of claim 23, wherein the expression level is normalized. The method of claim 23 or 24, wherein at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, or 15. The expression level of the dog marker gene is measured. The method of claim 25, wherein the expression levels of IFITM1, RGS13, CD79B, CD22, BTG2, CD44, EPDR1, and UAP1 are measured. The method of any one of claims 23-26, wherein the report comprises a recommendation for anti-CD40 antibody treatment for the subject. The method of claim 27, wherein the anti-CD40 antibody treatment is treatment with an agonist anti-CD40 antibody, and increased expression of one or more of IFITM1, CD79B, IGF1R, CD44, CTSC, EPDR1, and PUS7 relative to the reference level is And the subject is less likely to respond to agonist anti-CD40 antibody treatment. The method of claim 28, wherein the reference level is determined based on the expression level of the corresponding marker gene in a sample comprising B lymphoma cells from a subject having increased tumor volume after anti-CD40 antibody treatment. The method of claim 29, wherein the sample from the subject for determining the reference level comprises B lymphoma cells of the same type as the sample from the subject for which a personalized genomic profile is created. The method of claim 27, wherein the anti-CD40 antibody treatment is treatment with an agonist anti-CD40 antibody, and increased expression of one or more of CD40, RGS13, VNN2, LMO2, CD22, BTG2, and UAP1 relative to a reference level is Wherein the subject indicates whether it will respond to agonist anti-CD40 antibody treatment. The method of claim 31, wherein the reference level is determined based on the expression level of the corresponding marker gene in the sample comprising B lymphoma cells from a subject whose tumor volume has been reduced after anti-CD40 antibody treatment. 33. The method of claim 32, wherein the sample from the subject for determining the reference level comprises B lymphoma cells of the same type as the sample from the subject for which a personalized genomic profile is created. 34. The method of any one of claims 28-33, wherein the agonist anti-CD40 antibody stimulates CD40 and enhances the interaction between CD40 and CD40 ligand. The method of claim 34, wherein the agonist anti-CD40 antibody comprises the heavy chain amino acid sequence set forth in SEQ ID NO: 1 and the light chain amino acid sequence set forth in SEQ ID NO: 2. 34. The method of any one of claims 28-33, wherein the agonist anti-CD40 antibody stimulates CD40 and does not enhance or inhibit the interaction between CD40 and CD40 ligand. 37. The method of any one of claims 23-36, wherein the B cell lymphoma is diffuse large cell B-cell lymphoma (DLBCL). 37. The method of any one of claims 23-36, wherein the B cell lymphoma is a non-Hodgkin's lymphoma. The method of claim 38, wherein the non-Hodgkin's lymphoma is follicular lymphoma, mantle cell lymphoma, marginal lymphoma, or small lymphocytic lymphoma. 40. The method of any one of claims 23-39, wherein the sample comprising B lymphoma cells is a formalin fixed paraffin embedded biopsy sample. 41. The method of any one of claims 23-40, wherein the expression level of one or more marker genes is measured by the level of RNA transcript of one or more marker genes. The method of claim 41, wherein the RNA transcript is measured by qRT-PCR. 41. The method of any one of claims 23-40, wherein the expression level of one or more marker genes is measured by the level of protein expression of one or more marker genes. (a) at least 2 selected from the group consisting of IFITM1, CD40, RGS13, VNN2, LMO2, CD79B, CD22, BTG2, IGF1R, CD44, CTSC, EPDR1, UAP1, and PUS7 in a sample comprising B lymphoma cells from a subject Measuring expression levels of dog marker genes;
(b) calculating a sensitivity index (SI) value based on the measured expression level of the marker gene in step (a) by the following equation,

Wherein the expression levels of at least one marker gene having a positive correlation value and at least one marker gene having a negative correlation value are determined;
Wherein (i) β _j is the coefficient value for each marker gene measured; (ii) p is the number of marker genes measured; (iii) χ _i is a normalized expression level, transformed for a sample from the subject for the expression level of each marker measured; (iv) μ _j and σ _j are the mean and standard deviation for each marker gene measured; Wherein β _j , μ _j and σ _j are determined from a patient sample comprising B lymphoma cells from a clinical trial;
A value above 0 for the sensitivity index indicates whether the subject will respond to anti-CD40 antibody treatment, or a value below 0 for the sensitivity index indicates whether the subject will respond less to anti-CD40 antibody treatment. A method for predicting responsiveness of a subject with B-cell lymphoma to CD40 antibody treatment. The expression level of claim 44 wherein the expression level of at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, or 14 marker genes is measured and , Used for calculating the sensitivity index. The method of claim 44, wherein the expression levels of IFITM1, RGS13, CD79B, CD22, BTG2, CD44, EPDR1, and UAP1 are measured and used for calculating the sensitivity index. The method of claim 44, wherein β _j , μ _j and σ _j are determined from a patient sample having B lymphoma cells of the same type as the sample from the subject for which responsiveness to anti-CD40 treatment is predicted. (omission). At least one marker gene selected from the group consisting of IFITM1, CD79B, IGF1R, CD44, CTSC, EPDR1, PUS7, CD40, RGS13, VNN2, LMO2, CD22, BTG2, and UAP1 in a sample comprising B lymphoma cells from a subject Kit comprising a reagent for measuring the expression level of. The kit of claim 49, wherein the reagent comprises at least one pair of primers and probes for detecting the expression level of each marker gene by qRT-PCR. 51. The method of claim 49 or 50, further comprising instructions for assessing whether a human subject with B-cell lymphoma will respond to anti-CD40 antibody treatment based on the expression levels of one or more marker genes measured. Kit. 51. The method of claim 50, wherein the probe and primer pairs comprise SEQ ID NOs: 27, 28, and 29; SEQ ID NOs: 60, 61, and 62; SEQ ID NOs: 93, 94, and 95; SEQ ID NOs: 24, 25, and 26; SEQ ID NOs: 57, 58, and 59; SEQ ID NOs: 90, 91, and 92; SEQ ID NOs: 114, 115, and 116; SEQ ID NOs: 126, 127, and 128; SEQ ID NOs: 30, 31, and 32; SEQ ID NOs: 63, 64, and 65; SEQ ID NOs: 96, 97, and 98; SEQ ID NOs: 12, 13, and 14; SEQ ID NOs: 45, 46, and 47; SEQ ID NOs: 78, 79, and 80; SEQ ID NOs: 141, 142, and 143; SEQ ID NOs: 150, 151, and 152; SEQ ID NOs: 159, 160, and 161; SEQ ID NOs: 15, 16, and 17; SEQ ID NOs: 48, 49, and 50; SEQ ID NOs: 81, 82, and 83; SEQ ID NOs: 9, 10, and 11; SEQ ID NOs: 42, 43, and 44; SEQ ID NOs: 75, 76, and 77; SEQ ID NOs: 6, 7, and 8; SEQ ID NOs: 39, 40, and 41; SEQ ID NOs: 72, 73, and 74; SEQ ID NOs: 174, 175, and 176; SEQ ID NOs: 180, 181, and 182; SEQ ID NOs: 186, 187, and 188; SEQ ID NOs: 165, 166, and 167; SEQ ID NOs: 168, 169, and 170; SEQ ID NOs: 171, 172, and 173; SEQ ID NOs: 21, 22, and 23; SEQ ID NOs: 54, 55, and 56; SEQ ID NOs: 87, 88, and 89; SEQ ID NOs: 129, 130, and 131; SEQ ID NOs: 132, 133, and 134; SEQ ID NOs: 135, 136, and 137; SEQ ID NOs: 138, 139, and 140; SEQ ID NOs: 147, 148, and 149; SEQ ID NOs: 156, 157, and 158; SEQ ID NOs: 177, 178, and 179; SEQ ID NOs: 183, 184, and 185; And SEQ ID NOs: 189, 190, and 191. The kit of any one of claims 49-52, further comprising a reagent for measuring the expression level of BCL6 in a sample comprising B lymphoma cells from the subject. The kit of claim 53, wherein the reagent comprises at least one pair of primers and a probe to detect the expression level of BCL6 by qRT-PCR. 55. The kit of claim 54, wherein the probe and primer pairs are SEQ ID NO: 102, 103, and 104, or SEQ ID NO: 108, 109, and 110.

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